Drug-loaded biodegradable microbead compositions including drug-containing vesicular agents

ABSTRACT

Drug-loaded microbead compositions include microbeads of a biodegradable material and vesicular agents located within or associated with the biodegradable material of the microbeads. The vesicular agents include a lipid bilayer and comprise liposomes or ethosomes. The drug-loaded microbeads include a first therapeutic agent associated with the vesicular agents, and a second therapeutic agent different from the first therapeutic agent. The vesicular agents include a lipid bilayer surrounding a vesicular core. The second therapeutic agent is contained within the microbeads or associated with the microbeads through ionic or non-covalent interaction and may or may not be associated with the vesicular agents. Drug-loaded biodegradable microbead compositions include microbeads of biodegradable material and a therapeutic agent.

CROSS-REFERENCES TO RELATED APPLICATIONS

The present application is a continuation-in-part of U.S. applicationSer. No. 16/955,275, filed Jun. 18, 2020, which is a national-phaseapplication of International Application No. PCT/US2018/066232, filedDec. 18, 2018, which claims the benefit of priority to U.S. ProvisionalApplication Ser. No. 62/607,080, filed Dec. 18, 2017, the presentapplication being also a continuation-in-part under 35 U.S.C. §§ 111(a)of International Application No. PCT/US2019/037984 designating theUnited States, filed Jun. 19, 2019. All of the foregoing prioritydocuments are hereby incorporated by reference into this disclosure intheir entireties.

TECHNICAL FIELD

This disclosure relates to microbeads for chemoembolization, to methodsfor preparing the microbeads, to methods for using the microbeads, andto biodegradable drug-eluting particles containing therapeutic agentsfor chemoembolization.

BACKGROUND

Embolization therapy is a minimally invasive surgery performed byinterventional radiologists. Typical treatments may include entering thevasculature via a minor incision, such as in the arm or leg, and gainingaccess to the treatment site by use of guidewires and catheters,optionally aided by imaging techniques such as fluoroscopy. The embolicagent at the treatment site embolizes the vessel, blocking off the flowof blood to tumors downstream from the treatment site and resulting innecrosis and/or shrinkage of the tumors.

The embolic agent of choice for embolization therapy depends on thedesired clinical outcome, as well as the inherent properties of theembolic agent. Embolic agents used clinically suffer common drawbacks.First, the embolic agents are provided without preloaded drug, meaningthat the physician must order a pharmacy to load the drug into theembolic agent well in advance of surgery, typically at least 24 hoursbefore the surgery. Second, the embolic agents have a suboptimalpharmacokinetic release profile that stops releasing drug into thetarget treatment site after only a few days after implantation,typically within about 3 days after implantation.

Among the diseases for which embolization therapy may be effective ishepatocellular carcinoma (HCC). Hepatocellular carcinoma accounts for80% to 90% of all primary liver cancers. The mortality of hepatocellularcarcinoma is high, and treatment options are limited. Patients withcirrhosis exhibit a high probability of developing hepatocellularcarcinoma.

Sorafenib (co-developed and co-marketed by Bayer and OnyxPharmaceuticals as Nexavar) is an orally administered kinase inhibitordrug approved for the treatment of primary kidney cancer (advanced renalcell carcinoma), radioactive iodine resistant advanced thyroidcarcinoma, and advanced primary liver cancer such as hepatocellularcarcinoma. With the exception of sorafenib tosylate (the toluene salt ofsorafenib), no targeted agent is currently approved for the treatment ofhepatocellular carcinoma.

Cytotoxic agent doxorubicin exhibits a broad spectrum of antitumoractivity. When administrated via the hepatic artery, doxorubicinexhibits antitumor effects and partial response in hepatocellularcarcinoma patients. An exploratory analysis of a randomized phase IIstudy in hepatocellular carcinoma comparing doxorubicin alone todoxorubicin plus sorafenib showed a significant improvement in overallsurvival favoring the combination.

Ongoing needs exist for embolic agents and embolization therapy methodsthat enable greater efficiency in preparation of the embolic agents andthat also provide sustained, longer-term delivery of therapeutic agentto the embolization site.

Ongoing needs exist for embolic agents and embolization therapy methodsthat enable greater efficiency in preparation of the embolic agents andthat also provide sustained, longer-term delivery of therapeutic agentto the embolization site. Furthermore, ongoing needs exist for embolicagents that are capable of delivering multiple therapeutic agents,including but not limited to the combination of sorafenib anddoxorubicin, in a controlled manner as an alternative to oraladministration.

SUMMARY

Accordingly, embodiments of this disclosure include drug-loadedmicrobead compositions, methods for preparing the drug-loaded microbeadcompositions, embolization compositions prepared from the drug-loadedmicrobead compositions, methods for preparing the embolizationcompositions, and methods for treating diseases with the embolizationcompositions.

According to embodiments, a drug-loaded microbead composition mayinclude microbeads comprising a water-swellable polymer material; and acomplex comprising a carrier and a therapeutic agent chemically bondedto the carrier. In the drug-loaded microbead compositions, the complexis embedded within the polymer material. The drug-loaded microbeadcomposition have a water content of less than 1% by weight, based on thetotal weight of the drug-loaded microbead composition.

According to embodiments, methods for preparing a drug-loaded microbeadcomposition may include dissolving a carrier and a therapeutic agent inan aqueous solvent to form an initial mixture, the initial mixturecomprising a complex of the carrier and the therapeutic agent. Themethods may further include combining the initial mixture with a firstsolvent composition in which the therapeutic agent is less soluble thanin the aqueous solvent to form a second mixture, and stirring the secondmixture rapidly to form particles of the complex in the second mixture.The methods may further include combining the particles of the complexwith a hydrogel of a water-swellable polymer material to form a hydrogelmixture; combining the hydrogel mixture with a second solventcomposition to form a synthesis mixture; stirring the synthesis mixturerapidly to form drug-loaded microbeads in the synthesis mixture; andrecovering the drug-loaded microbeads from the synthesis mixture. Themethods may further include removing water from the drug-loadedmicrobeads to form a drug-loaded microbead composition having a watercontent of less than 1% by weight, based on the total weight of thedrug-loaded microbeads in the drug-loaded microbead composition.

According to embodiments, methods for preparing a drug-loaded microbeadcomposition may include dissolving a carrier and a therapeutic agent inan aqueous solvent to form an initial mixture, the initial mixturecomprising a complex of the carrier and the therapeutic agent; combiningthe initial mixture with a hydrogel solution of a water-swellablepolymer material to form a hydrogel mixture; combining the hydrogelmixture with a solvent composition to form a synthesis mixture; stirringthe synthesis mixture rapidly to form drug-loaded microbeads in thesynthesis mixture; and recovering the drug-loaded microbeads from thesynthesis mixture. The methods may further include drying thedrug-loaded microbeads to form a drug-loaded microbead compositionhaving a water content of less than 1% by weight, based on the totalweight of the drug-loaded microbeads in the drug-loaded microbeadcomposition.

According to embodiments, an embolization composition may include adrug-loaded microbead composition according to any embodiment of thisdisclosure, in combination with an amount of an aqueous solutionsufficient to cause the drug-loaded microbeads of the drug-loadedmicrobead composition to swell. In the embolization compositions, thedrug-loaded microbeads after swelling comprise a water content of from50% by weight to 95% by weight, based on the total weight of thedrug-loaded microbeads.

According to embodiments, methods for preparing an embolizationcomposition may include adding a drug-loaded microbead compositionaccording to any embodiment of this disclosure, or a drug-loadedmicrobead composition made according to the method of preparing thedrug-loaded microbead composition according any embodiment of thisdisclosure, to an amount of an aqueous solution sufficient to cause thedrug-loaded microbeads of the drug-loaded microbead composition to swelland form an injection-ready solution. In the embolization solution madeaccording to such methods, the drug-loaded microbeads after swellinghave a water content of from 50% by weight to 95% by weight, based onthe total weight of the drug-loaded microbeads. The methods may furtherinclude loading the injection-ready solution into an injection device.

According to embodiments, methods for treating a disease may includeproviding intravenously, to a subject in need of embolization therapy,an embolization composition according to any embodiment of thisdisclosure.

Further embodiments are directed to drug-loaded microbead compositionsincluding a plurality of individual microbeads of a biodegradablematerial, a plurality of individual vesicular agents, a firsttherapeutic agent, and a second therapeutic agent different from thefirst therapeutic agent. The vesicular agents include at least one lipidbilayer surrounding a vesicular core. The first therapeutic agent isassociated with the individual vesicular agents. For example, the firsttherapeutic agent may be contained within the individual vesicularagents or may be associated with the individual vesicular agents byionic or non-covalent interaction. The second therapeutic agent isdifferent from the first therapeutic agent, is contained within theindividual microbeads or associated with the individual microbeadsthrough ionic or non-covalent interaction, and may or may not beassociated with the individual vesicular agents within the individualmicrobeads.

Example embodiments disclosed herein are directed to methods forpreparing the drug-loaded microbead compositions, to embolizationcompositions prepared from the drug-loaded microbead compositions, tomethods for preparing the embolization compositions, and to methods fortreating a disease using the embolization compositions.

Further embodiments of this disclosure are directed to drug-loadedbiodegradable microbead compositions. The drug-loaded biodegradablemicrobead compositions may include microbeads of a biodegradablematerial and at least one therapeutic agent embedded in a matrix of thebiodegradable material, encapsulated by a matrix of the biodegradablematerial, disposed within a porous structure formed by the matrix of thebiodegradable material, or ionically or non-covalently associated with amatrix of the biodegradable material. In such embodiments of drug-loadedbiodegradable microbead compositions, the microbeads of thebiodegradable material may contain, but need not necessarily contain,vesicular agents such as liposomes or ethosomes.

BRIEF DESCRIPTION OF THE DRAWING

The FIGURE is a graph of in vivo drug release vs. time for (A)drug-loaded microbeads in an embolization composition according to oneor more embodiments of this disclosure; (B) a comparative embolizationcomposition containing microbeads of biocompatible sulfonate-modifiedpoly(vinyl alcohol) (PVA) loaded with doxorubicin; and (C) a comparativeembolization composition containing microbeads of a sodium acrylatealcohol copolymer formed by polymerization of vinyl acetate and methylacrylate and loaded with doxorubicin.

Additional features and advantages of the embodiments described hereinwill be set forth in the detailed description which follows, and in partwill be readily apparent to those skilled in the art from thatdescription or recognized by practicing the embodiments describedherein, including the detailed description which follows, the claims, aswell as the appended drawings.

It is to be understood that both the foregoing general description andthe following detailed description describe various embodiments and areintended to provide an overview or framework for understanding thenature and character of the claimed subject matter. The accompanyingdrawings are included to provide a further understanding of the variousembodiments, and are incorporated into and constitute a part of thisspecification. The drawings illustrate the various embodiments describedherein, and together with the description serve to explain theprinciples and operations of the claimed subject matter.

DETAILED DESCRIPTION

Some embodiments of this disclosure are directed to drug-loadedmicrobead compositions, to methods for preparing the drug-loadedmicrobead compositions, to embolization compositions prepared from thedrug-loaded microbead compositions, to methods for preparing theembolization compositions, and to methods for treating a disease usingthe embolization compositions. The drug-loaded microbead compositionsinclude microbeads into which a therapeutic agent has been pre-loaded.When included in an embolization composition and provided to a subjectas part of an embolization therapy, the embolization compositions mayprovide a continuous release of the drug into the target treatment sitefor at least 7 days, at least 14 days, at least 21 days, or at least 30days after implantation.

A drug-loaded microbead composition may include microbeads loaded with atherapeutic agent or with a complex of a therapeutic agent and acarrier. The microbeads may be beads or microparticles of awater-swellable polymer material, such as a polymer material capable offorming a hydrogel.

In the microbeads of the drug-loaded microbead composition, thetherapeutic agent or complex of the therapeutic agent may be embeddedwithin the water-swellable polymer materials. Except when describedotherwise, the term “embedded” broadly includes embodiments for whichthe water-swellable polymer material or some portion thereof generallysurrounds the therapeutic agent or the complex including the therapeuticagent or is embedded by the therapeutic agent or the complex includingthe therapeutic agent. For example, “embedded” may include in someembodiments a water-soluble polymer material shell that encapsulates acore that holds the therapeutic agent or complex. In other embodiments,“embedded” may include a structure in which the therapeutic agent or thecomplex is physically disposed within a matrix, network, or porestructure of a water-swellable polymer material that may or may not havea core within an outer shell. The therapeutic agent itself is notchemically bonded to the water-swellable polymer material at all or isnot chemically bonded directly to the polymer backbone of thewater-swellable polymer material. The drug-loaded microbead compositionmay have a water content of less than 1% by weight, based on the totalweight of the drug-loaded microbead composition.

In various embodiments, the water-swellable polymer material of thedrug-loaded microbead composition may include a natural hydrogel polymersuch as a chitosan or a polysaccharide, or a synthetic hydrogel polymersuch as a polyacrylate, a polyamide, a polyester, a polysaccharide, apoly(methylmethacrylate), or a poly(vinyl alcohol), for example. In someembodiments, the water-swellable polymer material may be biodegradable.Specific examples of water-swellable polymer materials include, withoutlimitation, poly(-hydroxybutyrate), methacrylated hyaluronic acids(hyaluronic acids being polymers of disaccharides composed ofD-glucuronic acid and N-acetyl-D-glucosamine), chitosan-alginates,poly(N-isopropylacrylamide) copolymers,poly(N-isopropylacrylamide)-alginates,poly(N-isopropylacrylamide)-peptides,poly(N-isopropylacrylamide)-α-acryloyloxy-β,β-dimethyl-γ-butyrolactone-hydrophilicJeffamine, or poly(N-isopropyl-acrylamide)-poly(ethylene glycol)diacrylate-pentaerythritol tetrakis(3-mercapto-propionate). Thewater-swellable polymer materials of the drug-loaded microbeadcomposition may include derivatives of any of the foregoing materials,or may include combinations of any of the foregoing materials or theirderivatives. For example, the drug-loaded microbead compositions mayinclude a combination of multiple water-swellable polymer materials, inwhich each individual microbead is made of a single type of polymer, andthe drug-loaded microbead composition includes microbeads of multiplepolymer types. Alternatively, the drug-loaded microbead compositions mayinclude a combination of multiple water-swellable polymer materials, inwhich individual microbeads of the composition are composed of multipletypes of polymer.

The drug-loaded microbead composition includes a therapeutic agent. Insome embodiments, the therapeutic agent may be a hydrophilic therapeuticagent or a therapeutic agent that either is water soluble or has atleast some solubility in an aqueous solution. In some embodiments, thetherapeutic agent may be a chemotherapeutic agent having at least someefficacy for treating a disease such as cancer. In some embodiments, thetherapeutic agent may be a chemotherapeutic agent having at least someefficacy for treating a cancer such as hepatocellular carcinoma, livercancer, prostate cancer, or breast cancer. The therapeutic agent mayhave one or more chemical moieties or atomic centers having a positiveor negative charge or affinity. Examples of specific therapeutic agentsinclude, without limitation, doxorubicin, sorafenib, vandetanib,nivolumab, ipilimumab, regorafenib, irinotecan, epirubicin, pirarubicin,5-fluorouracil, cisplatin, floxuridine, mitomycin C, derivatives of anyof the foregoing, prodrugs of any of the foregoing, therapeuticallyacceptable salts or crystalline forms of any of the foregoing, orcombinations of any of the foregoing. Further examples of suitabletherapeutic agents include, without limitation, pirarubicin,mitoxantrone, tepotecan, paclitaxel, carboplatin, pemetrexed,penistatin, pertuzumab, trastuzumab, and docetaxel.

Individual microbeads of the drug-loaded microbead composition mayinclude one therapeutic agent or a plurality of therapeutic agents.Collectively, the microbeads of the drug-loaded microbead compositionmay include some microbeads loaded with one specific therapeutic agentor combination of specific therapeutic agents and other microbeadsloaded with a different specific therapeutic agent or combination ofspecific therapeutic agents.

In some embodiments, the therapeutic agent of the drug-loaded microbeadcomposition may be embedded within the microbeads of the water-swellablepolymer material but not chemically bonded to the water-swellablepolymer material. In some embodiments, the therapeutic agent of thedrug-loaded microbead composition may be embedded within the microbeadsof the water-swellable polymer material and not chemically bondeddirectly to the polymer backbone water-swellable polymer material, yetmay be chemically bonded to a functional group of the water-swellablepolymer material. As used herein, “not chemically bonded” refers to alack of covalent chemical bonds between the therapeutic agent and thewater-swellable polymer but does not preclude the existence ofnoncovalent intermolecular interactions such as ionic interactions or avan der Waals interaction between the therapeutic agent and thewater-swellable polymer material.

In some embodiments, the drug-loaded microbead composition includes acomplex of a carrier and a therapeutic agent. In the complex, thetherapeutic agent may be chemically bonded to the carrier or may beassociated with the carrier by a non-covalent means such asencapsulation or a van der Waals interaction. The complex may beembedded within the water-swellable polymer material. When the complexis embedded within the microbeads, the carrier may be chemically bondedto the water-swellable polymer material while the therapeutic agent isnot chemically bonded to the water-swellable polymer material. Withoutintent to be bound by theory, it is believed that when the therapeuticagent is bonded or associated with the carrier but is not chemicallybonded to the water-swellable polymer material, the microbeads of thedrug-loaded microbead composition are less susceptible to shrinking as aresult of replacing water molecules with drug molecules during drugloading. Accordingly, the final size distribution of the drug-loadedmicrobeads can be controlled more readily by selecting appropriatemicrobead sizes before the therapeutic agent is loaded.

When the drug-loaded microbead composition includes a complex of thecarrier and the therapeutic agent, the carrier may be anypharmaceutically acceptable compound that can complex with orencapsulate the therapeutic agent. In some embodiments, the carrier mayhave charged chemical groups or chemical groups with dipole moments thatinteract with corresponding chemical groups of the therapeutic agenthaving an opposite charge or opposite dipole moment. If the carrier is apolymeric material, the carrier is a different material from thewater-swellable polymer material. Non-limiting examples of suitablecarriers include polysaccharides, liposomes, polymeric micelles,Pluronics, polycaprolactone-b-methoxy-PEG, poly(aspartic acid)-b-PEG,poly(benzyl-L-glutamate)-b-PEG, poly(D,L-lactide)-b-methoxy-PEG,poly(β-benzyl-L-asparate)-b-PEG). Non-limiting examples ofpolysaccharides include dextrans and dextran sulfates such as dextransodium sulfate. In one example embodiment, the carrier may include adextran sodium sulfate having a weight-average molecular weight of fromabout 40 kDa (kilodalton) to about 500 kDa, or from about 50 kDa toabout 300 kDa, or from about 100 kDa to about 300 kDa, or about 100 kDato about 200 kDa.

The drug-loaded microbead compositions according to embodiments may havea very low water content such as less than 1%, or less than 0.1%, orless than 0.05% (500 ppm), or less than 0.02% (200 ppm), or less than0.01% (100 ppm), or less than 0.005 (50 ppm), or less than 0.002% (20ppm), or less than 0.001% (10 ppm) by weight water, based on the totalweight of the microbeads in the drug-loaded microbead composition.Without intent to be bound by theory, it is believed that a very lowwater content of the drug-loaded microbead composition increases theshelf-life and long-term stability of the drug-loaded microbeadcomposition. Further, it is believed that water contents significantlygreater than 1% by weight (such as 2%, 3%, 5%, or 10%, for example)based on the total weight of the drug-loaded microbead composition, maylead to decomposition or hydrolysis of the therapeutic agent,instability or breaking apart of the water-swellable polymer, or acombination of these, within a few days or even a few hours, such thatthe composition cannot be used for embolization procedures, even if thedrug-loaded microbead composition is rehydrated. It is believed that theshelf-life and long-term stability of compositions having water contentssignificantly greater than 1% by weight are not sufficiently long toensure viability of the therapeutic agent over the time period frommanufacture of the drug-loaded microbead composition to use of thecomposition in an embolization procedure. It is believed that selectionof the water-swellable polymer material may correlate with the abilityfor water to be removed from the drug-loaded microbeads bylyophilization or other drying technique or combination of dryingtechniques in an amount sufficient to prevent decomposition of thetherapeutic agent.

A very low water content of the drug-loaded microbead composition, aspreviously described may be attained by drying techniques that will bedescribed in greater detail subsequently, with respect to methods forpreparing the drug-loaded microbead compositions. In this regard, thedrug-loaded microbead compositions may be dry or nearly dehydratedcompositions of the microbeads containing the embedded therapeutic agentor the embedded complex of the therapeutic agent and the carrier. Thedrug-loaded microbead compositions may have a powder-like consistency.Accordingly, the drug-loaded microbead compositions may be made suitablefor injection into a subject being treated by rehydrating the microbeadsof the drug-loaded microbead compositions to form an embolizationcomposition, as will be described in greater detail subsequently.Regardless, the drug-loaded microbead compositions may be provided insuch a form that a physician needs to add only an aqueous solution suchas water or physiologically buffered saline solution to the drug-loadedmicrobead composition to prepare the composition for use in anembolization procedure.

The microbeads of the drug-loaded microbead composition may have anyshape common to microparticles formed from a hydrogel typewater-swellable polymer material. For example, the microparticles may bespherical or substantially spherical, or may have an ovoid shape withoval-shaped or elliptical cross-sections about a longitudinal axis andcircular cross-sections about an axis perpendicular to the longitudinalaxis.

The drug-loaded microbead composition may include an amount oftherapeutic agent per unit volume of microbeads in the composition thatis chosen to have a desired therapeutic effect or activity, based on theintended use for the drug-loaded microbead composition and theparticular therapeutic agent present in the individual microbeads.

The amount of therapeutic agent in the individual microbeads of thedrug-loaded microbead composition can be adjusted through particulartechniques involved during drug loading, such as loading time, loadingtemperature, or concentration of therapeutic agent in a loadingsolution, for example. The amount of therapeutic agent in the individualmicrobeads of the drug-loaded microbead composition can be adjusted alsothrough synthetic techniques involved for synthesizing the microbeadsthemselves, such as through adjusting polymer molecular weights, degreeof hydrogel crosslinking, polymer density, or polymer porosity of thewater-swellable polymer material. For example, when doxorubicin is thetherapeutic agent, the amount of drug loading in the drug-loadedmicrobeads can be adjusted with respect to the number of negativecharges in the polymer backbone of the water-swellable polymer material.Similarly, when sorafenib is the therapeutic agent, the sorafenib may beembedded within polymeric micelles or liposomes that can be embeddedwithin the microbead structure. The amount of therapeutic agent in theindividual microbeads of the drug-loaded microbead composition can beadjusted also through choice of the carrier.

In example embodiments, the individual drug-loaded microbeads of thedrug-loaded microbead composition include the water-swellable polymermaterial, the therapeutic agent, the carrier, and water. The individualdrug-loaded microbeads of the drug-loaded microbead composition mayinclude from about 30% by weight to about 70% by weight, or from about35% by weight to about 65% by weight, or from about 40% to about 60% byweight, or about 45% by weight to about 55% by weight, or about 50% toabout 70% by weight water-swellable polymer material, based on the totalweight of the individual drug-loaded microbead. In example embodiments,the individual drug-loaded microbeads of the drug-loaded microbeadcomposition may include from about 1% by weight to about 25% by weight,or from about 1% by weight to about 20% by weight, or from about 1% byweight to about 15% by weight, or from about 2% by weight to about 25%by weight, or from about 5% by weight to about 25% by weight, or fromabout 10% by weight to about 25% by weight therapeutic agent, based onthe total weight of the individual drug-loaded microbead. In exampleembodiments, the individual drug-loaded microbeads of the drug-loadedmicrobead composition may include from about 1% by weight to about 40%by weight, or from about 1% by weight to about 30% by weight, or fromabout 1% by weight to about 25% by weight, or from about 1% by weight toabout 20% by weight, or from about 5% by weight to about 40% by weight,or from about 10% by weight to about 40% by weight, or from about 20% byweight to about 40% by weight carrier, based on the total weight of theindividual drug-loaded microbead. In example embodiments, the individualdrug-loaded microbeads of the drug-loaded microbead compositionaccording to embodiments may have a very low water content such as lessthan 1% by weight, or less than 0.5% by weight, or less than 0.1% byweight, or less than 0.05% (500 ppm) by weight, or less than 0.02% (200ppm) by weight, or less than 0.01% (100 ppm) by weight, or less than0.005 (50 ppm) by weight, or less than 0.002% (20 ppm) by weight, orless than 0.001% (10 ppm) by weight water, based on the total weight ofthe individual microbeads in the drug-loaded microbead composition.

Having described drug-loaded microbead compositions according to variousembodiments, methods for preparing the drug-loaded microbeadcompositions will now be described.

In some embodiments, methods for preparing a drug-loaded microbeadcomposition as described previously may include dissolving a carrier anda therapeutic agent in an aqueous solvent to form an initial mixture.Within the initial mixture, complexes may form between molecules of thecarrier and molecules of the therapeutic agent. For example, a mixtureof doxorubicin as a therapeutic agent with a dextran sulfate as acarrier may form complexes of doxorubicin and dextran sulfate.

The initial mixture including the complexes may then be combined with afirst solvent composition to form a second mixture. The first solventcomposition includes a solvent or a mixture of solvents, in thecombination of which the therapeutic agent is less soluble than in theaqueous solvent. The second mixture may be a biphasic or a multiphasicmixture. Then, the second mixture is rapidly agitated or stirred. Therapid agitation or stirring causes particles of the complex to form inthe second mixture. The second mixture may be stirred, for example, at astirring speed from about 150 rpm to about 2000 rpm. The stirring may becontinuous and may be carried out for an amount of time sufficient toform microbeads. For example, the stirring may be carried outcontinuously for a stirring time from about 1 hour to about 12 hours. Ifthe second mixture is biphasic, the particles may form in the phase ofthe second mixture in which the therapeutic agent is less soluble orleast soluble. Optionally, the particles of the complex may be isolatedfrom the second mixture by any suitable technique such as filtration,for example.

The particles of the complex, either still in the second mixture oroptionally after isolation from the second mixture, may then be combinedwith a hydrogel of a water-swellable polymer material to form a hydrogelmixture. The hydrogel mixture is then combined with a second solventcomposition to form a synthesis mixture. The second solvent compositionincludes a solvent or a mixture of solvents, in the combination of whichthe therapeutic agent and/or the hydrogel is less soluble than inaqueous solvent.

The synthesis mixture is agitated or stirred rapidly to form drug-loadedmicrobeads in the synthesis mixture. The synthesis mixture may bestirred, for example, at a stirring speed from about 150 rpm to about2000 rpm. The stirring may be continuous and may be carried out for anamount of time sufficient to form microbeads. For example, the stirringmay be carried out continuously for a stirring time from about 1 hour toabout 12 hours. The drug-loaded microbeads may form either as thewater-swellable polymer cures around the complex of the therapeuticagent or as the complex of the therapeutic agent absorbs into thepolymer matrix of the water-swellable polymer. The drug-loadedmicrobeads are then recovered from the synthesis mixture by any suitableseparation technique such as filtration, for example.

The drug-loaded microbeads prepared according to the foregoing proceduremay exhibit a core-shell structure, in which a polymer matrix shellencapsulates a particle core and the complex is disposed in the particlecore. In the core-shell structure, the therapeutic agent is notchemically bonded to the water-swellable polymer material and thecarrier may be chemically bonded to the water-swellable polymermaterial.

In other embodiments, methods for preparing a drug-loaded microbeadcomposition as described previously may include dissolving a carrier anda therapeutic agent in an aqueous solvent to form an initial mixture.Within the initial mixture, complexes may form between molecules of thecarrier and molecules of the therapeutic agent. For example, a mixtureof doxorubicin as a therapeutic agent with a dextran sulfate as acarrier may form complexes of doxorubicin and dextran sulfate.Subsequently, the initial mixture may be combined with a hydrogelsolution of a water-swellable polymer material to form a hydrogelmixture, without first forming particles of the complex as anintermediate step.

The hydrogel mixture is then combined with a second solvent compositionto form a synthesis mixture. The second solvent composition includes asolvent or a mixture of solvents, in the combination of which thetherapeutic agent and/or the hydrogel is less soluble than in aqueoussolvent. The synthesis mixture is stirred rapidly for form drug-loadedmicrobeads in the synthesis mixture. The drug-loaded microbeads may formeither as the water-swellable polymer cures in the presence of thecomplex of the therapeutic agent or as the complex of the therapeuticagent absorbs into the polymer matrix of the water-swellable polymer.The drug-loaded microbeads are then recovered from the synthesis mixtureby any suitable separation technique such as filtration, for example.The drug-loaded microbeads prepared according to the foregoing proceduremay exhibit an interpenetrating network structure, in which thewater-swellable polymer material has a polymer matrix and the complexesof the therapeutic agent and the carrier are disposed within porestructures of the polymer matrix. In the interpenetrating networkstructure, the therapeutic agent may be chemically bonded to thecarrier, and the complex of the therapeutic agent and the carrier may bedispersed within molecules of the water-swellable polymer.

The foregoing methods for preparing drug-loaded microbead compositions,regardless of whether the drug-loaded microbeads are prepared includingthe intermediate step of forming particles of the complexes oftherapeutic agent and carrier or without the intermediate step, furtherinclude removing water from the drug-loaded microbeads to form adrug-loaded microbead composition having a water content of less than 1%by weight, based on the total weight of the drug-loaded microbeadcomposition. In some embodiments, the removal of the water may includelyophilization or freeze drying, optionally followed by an additionaldrying step involving temperature variation (heating or cooling),flowing air, vacuum, or a combination of these. The drug-loadedmicrobeads may have an average synthesized volume when recovered fromthe synthesis mixture and an average final volume after the removing ofwater. The average final volume may range from about 10% to about 75% ofthe average synthesized volume such as, for example, about 10%, about20%, about 30%, about 40%, about 50%, about 60%, about 70%, or about 75%of the average synthesized volume.

Lyophilization or freeze drying of the drug-loaded microbeads may beconducted according to any procedure commonly used for drying ofparticulate substances. For example, the drug-loaded microbeads may beplaced in partially stoppered glass vials, which then are placed on acooled, temperature-controlled shelf within a freeze dryer. The shelftemperature is reduced, and the sample is frozen to a uniform, definedtemperature. After complete freezing, the pressure in the dryer may bedecreased to a defined pressure to initiate primary drying. During theprimary drying, water vapor is progressively removed from the frozenmass by sublimation while the shelf temperature is controlled at aconstant, low temperature. Secondary drying may be initiated byincreasing the shelf temperature and reducing the chamber pressurefurther so that water absorbed to the semi-dried mass can be removeduntil the residual water content decreases to the desired level.

Lyophilization or freeze drying of the drug-loaded microbeads may alsobe conducted by atmospheric pressure freeze drying by rapidlycirculating very dry air over the frozen drug-loaded microbeads. Thecirculating dry gas provides improved heat and mass transfer from thefrozen microbeads. Atmospheric spray drying processes may facilitate theformation of small-diameter microbeads as a free-flowing powder, whileavoiding formation of a dried cake. The free-flowing powder, in turn,may facilitate rehydration of the drug-loaded microbead composition whenan embolization composition is prepared.

The drug-loaded microbeads of the drug-loaded microbead compositionspreviously described, or prepared according to the foregoing methods,may exhibit physical or mechanical properties beneficial to their use,storage, transport, or subsequent rehydration to form an embolizationcomposition.

For example, it has been described previously that in the drug-loadedmicrobead compositions and embolization compositions according toembodiments of this disclosure, the therapeutic agent in the microbeadsis not chemically bonded to the water-swellable polymer. Without intentto be bound by theory, it is believed that the lack of chemical bondingdirectly between the therapeutic agent and the water-swellable polymercauses microbeads of the water-swellable polymer to have substantiallyidentical swollen diameters when loaded with therapeutic agent and whennot loaded with therapeutic agent, whether before therapeutic agent isloaded into the microbeads or after therapeutic agent is eluted out ofthe microbeads.

In contrast, it has been observed that, some hydrogel polymer microbeadsnot according to embodiments of this disclosure have, in their swollenstate without drug loading, an original diameter that shrinks by as muchas 5% or 10% when drug is loaded into the polymer matrix of themicrobeads. Without intent to be bound by theory, it is believed thatsuch shrinkage results because the addition of drug molecules to themicrobeads causes chemical bonds between the drug and the microbeadpolymer that, in turn, limit the amount of water that can be present inthe microbead polymer to swell the microbead polymer.

In clinical practice, the shrinkage observed in some hydrogel polymermicrobeads not according to embodiments of this disclosure is reversedwhen such a microbead composition is prepared for injection into apatient, because some amount of drug elutes out of the microbeads duringthe preparation process. Reversal of the shrinkage is exhibited as anincrease in the diameters of the microbeads, even though drug is beingremoved or eluted. The diameters of such may become sufficiently largeto clog a microcatheter being used to inject the microbeads. It isbelieved that the observed issues of microcatheter clogging are avoidedthrough the drug-loaded microbead compositions and embolizationcompositions according to embodiments of this disclosure, in which thetherapeutic agent is not chemically bonded to the water-swellablepolymer material.

As a further illustration, mechanical properties of the drug-loadedmicrobeads may be tailored by adjusting properties of, or syntheticmethods for, the water-swellable polymer material. For example,compression strength of the drug-loaded microbeads may be adjusted bychanging the crosslinking agents used to prepare the microbeads or bychanging the density or molecular weight of the water-swellable polymermaterials through polymer synthesis techniques known and understood bythose skilled in the art of polymer synthesis. In some embodiments, thedrug-loaded microbeads of the drug-loaded microbead compositions mayhave physical and mechanical properties chosen, such that thedrug-loaded microbeads recover greater than 85% of their diameter afterbeing compressed to 50% of their initial diameter.

The drug-loaded microbeads of the drug-loaded microbead compositionspreviously described, or prepared according to the foregoing methods,may be provided to a physician with instructions for rehydrating thedrug-loaded microbead composition to form an embolization compositionthat is ready for use in an embolization treatment. Accordingly,embodiments of this disclosure include embolization compositions andmethods for preparing the embolization compositions.

An embolization composition may include a drug-loaded microbeadcomposition according to any embodiment of this disclosure or adrug-loaded microbead composition prepared according to an embodiedmethod of this disclosure. The embolization composition may furtherinclude amount of an aqueous solution sufficient to cause thedrug-loaded microbeads of the drug-loaded microbead composition toswell. In particular, the amount of aqueous solution may cause thedrug-loaded microbeads, after swelling, to have a water content of fromabout 50% by weight to about 99% by weight, from about 60% by weight toabout 95% by weight, from about 70% by weight to about 95% by weight,from about 80% by weight to about 95% by weight, from about 90% byweight to about 95% by weight, or from about 80% by weight to about 99%by weight, or from about 90% by weight to about 99% by weight, or fromabout 95% by weight to about 99% by weight, based on the total weight ofthe drug-loaded microbeads. The aqueous solution may be anypharmaceutically acceptable solution such as, for, example, aphysiologically buffered saline solution.

In example embodiments, the swollen individual drug-loaded microbeads ofthe embolization composition include the water-swellable polymermaterial, the therapeutic agent, the carrier, and water. The swollenindividual drug-loaded microbeads of the embolization composition mayinclude from about 3% by weight to about 10% by weight, from about 3% byweight to about 7% by weight, or from about 3.5% by weight to about 6.5%by weight, or from about 4% to about 6% by weight, or about 4.5% byweight to about 5.5% by weight, or about 5% to about 7% by weightwater-swellable polymer material, based on the total weight of theswollen individual drug-loaded microbead. In example embodiments, theswollen individual drug-loaded microbeads of the embolizationcomposition may include from about 0.1% by weight to about 2.5% byweight, or from about 0.1% by weight to about 2% by weight, or fromabout 0.1% by weight to about 1.5% by weight, or from about 0.2% byweight to about 2.5% by weight, or from about 0.5% by weight to about2.5% by weight, or from about 1% by weight to about 2.5% by weighttherapeutic agent, based on the total weight of the swollen individualdrug-loaded microbead. In example embodiments, the swollen individualdrug-loaded microbeads of the embolization composition may include fromabout 0.1% by weight to about 4% by weight, or from about 0.1% by weightto about 3% by weight, or from about 0.1% by weight to about 2.5% byweight, or from about 0.1% by weight to about 2% by weight, or fromabout 0.5% by weight to about 4% by weight, or from about 1% by weightto about 4% by weight, or from about 2% by weight to about 4% by weightcarrier, based on the total weight of the swollen individual drug-loadedmicrobead. In example embodiments, the swollen individual drug-loadedmicrobeads of the embolization composition according to embodiments mayhave a water content of from about 50% by weight to about 99% by weight,from about 50% by weight to about 97% by weight, from about 50% byweight to about 96% by weight, from about 60% by weight to about 95% byweight, from about 70% by weight to about 95% by weight, from about 80%by weight to about 95% by weight, from about 90% by weight to about 95%by weight, or from about 80% by weight to about 99% by weight, or fromabout 90% by weight to about 99% by weight, or from about 95% by weightto about 99% by weight, based on the total weight of the individualdrug-loaded microbeads.

The swollen microbeads of the embolization composition may have averagediameters from about 5 μm to about 1200 μm, as measured by dynamic lightscattering (DLS). For non-spherical microbeads, the diameter of anindividual particle is taken to be the widest measurement attainablefrom a first point on the surface of the microbead, through the centerof mass of the microbead, to a second point on the surface of themicrobead opposite the first point. The specific size distribution ofthe microparticles may be chosen or tailored to suit the particulartreatment for which the embolization composition is intended to be used.The average diameters of the microbeads in the embolization compositionmay be chosen by any suitable method of size selection before or afterthe therapeutic agent is loaded into the microbeads. In someembodiments, the swollen microbeads of the embolization composition mayhave average diameters such as, for example, from about 40 μm to about800 μm, from about 40 μm to about 100 μm, from about 40 μm to about 75μm, from about 75 μm to about 100 μm, from about 100 μm to about 200 μm,from about 200 μm to about 300 μm, from about 300 μm to about 400 μm,from about 400 μm to about 500 μm, from about 600 μm to about 700 μm, orany subset of any of the foregoing ranges. In some embodiments, theswollen microbeads may have average diameters with a narrow sizedistribution such as 40 μm±20 μm, or 40 μm±10 μm, or 40 μm±5 μm, or 40μm±1 μm, for example.

Methods for preparing an embolization composition may include adding adrug-loaded microbead composition according to any embodiment of thisdisclosure, or a drug-loaded microbead composition prepared according toan embodied method of this disclosure, to an amount of an aqueoussolution sufficient to cause the drug-loaded microbeads of thedrug-loaded microbead composition to swell, whereby the drug-loadedmicrobeads after swelling comprise a water content of from 50% to 99% byweight, from 50% to 90% by weight, from 50% to 75% by weight, from 60%to 99% by weight, from 75% to 99% by weight, from 75% to 95% by weight,from 75% to 90% by weight, or from 85% to 99% by weight, based on thetotal weight of the drug-loaded microbeads, to form an injection-readysolution. The aqueous solution may be any pharmaceutically acceptablesolution such as, for, example, a physiologically buffered salinesolution. The methods may further include loading the injection-readysolution into an injection device such as a syringe, for example. Themethods may further include allowing the drug-loaded microbeads to swellfor a rehydration time of from about 5 minutes to about 60 minutesbefore loading the injection-ready solution into the injection device.In some examples, the embolization composition may contain from about 25mg therapeutic agent per milliliter of microbeads to about 150 mgtherapeutic agent per milliliter of microbeads.

The embolization compositions prepared from the drug-loaded microbeadcompositions according to embodiments of this disclosure may beincorporated into an embolization treatment or therapy intended to treata disease such as a cancer. In some embodiments, the therapeutic agentmay be a chemotherapeutic agent having at least some efficacy fortreating a cancer such as hepatocellular carcinoma, liver cancer,prostate cancer, or breast cancer. Accordingly, embodiments of thisdisclosure include methods for treating a disease. Methods for treatinga disease may include providing intravenously, to a subject in need ofembolization therapy, an embolization composition according to anyembodiment previously described herein, prepared from a drug-loadedmicrobead composition according to any embodiment previously describedherein.

In the methods for treating a disease, after the embolizationcomposition is provided intravenously, at least a portion of thedrug-loaded microbeads flow through vasculature of the subject to anembolization site and restrict blood flow through the embolization site.Thereafter, the drug-loaded microbeads may release to tissue at theembolization site over the course of a release period at least 90% byweight of the therapeutic agent, based on an initial amount oftherapeutic agent embedded within the microbeads before the embolizationcomposition is provided. In some embodiments, the release to the tissueat the embolization site may include an initial burst of drug release,during which at least 10% by weight of the therapeutic agent initiallypresent in the drug-loaded microbeads is released into surroundingtissues at the embolization site. The initial burst may occur within 1minute, 5 minutes, 10 minutes, 30 minutes, 60 minutes, 120 minutes, 180minutes, 240 minutes, 300 minutes, 360 minutes, 12 hours, 18 hours, or24 hours after the drug-loaded microbeads reach the embolization site.In some embodiments, the drug-loaded microbeads provide a sustainedrelease of the therapeutic agent to the tissue over the release period.The release period may be an extended release period such as at least 5days, at least 10 days, at least 14 days, at least 28 days, or at least42 days. The end of release period is determined from the time when thedrug-loaded microbeads are either completely decomposed or stop elutingtherapeutic agent to the tissue.

Without intent to be bound by theory, it is believed that the releasemechanism of the therapeutic agent from the drug-loaded microbeads ofthe embolization composition may be based on a dual mechanism includingan ion exchange process and an enzymatic process. For example, it isbelieved that in an early stage of delivery of therapeutic agent to theembolization site, when plenty of water is present, the therapeuticagent may be released from the drug-loaded microbeads by ion exchangeprocess. In a later stage, when water is lacking due to the vessel'sbeing embolized, the drug-loaded microbeads may be degraded by anenzymatic process such as by lysozymes, for example, to release thetherapeutic agent from the microbeads. It is believed that in the lateststage, such as by day 21 following implantation, most of thewater-swellable polymer material has been resorbed through tissuesurrounding the embolization site. Because at this point thewater-swellable material no longer has a matrix encapsulating thetherapeutic agent or the complex of the therapeutic agent and thecarrier, the therapeutic agent can be completely eluted into theembolization site. Thereby, it is believed that the embolizationcompositions according to embodiments of this disclosure may increasetumor response and free survival rates. Additionally, the embolizationcompositions according to embodiments of this disclosure provide aneconomic value such as reducing the physician procedure time byeliminating a need to request a pharmacist or technician to addtherapeutic agent to unloaded microbeads.

Reference will now be made in detail to embodiments of drug-loadedmicrobead compositions including microbeads or a biodegradable materialloaded with at least two therapeutic agents. The microbead compositionsaccording to embodiments include individual microbeads that contain aplurality of individual vesicular agents such as liposomes or ethosomes.At least one of the therapeutic agents is contained within the liposomesor ethosomes, either incorporated within a lipid bilayer or surroundedby a lipid bilayer. Another of the therapeutic agents may be containedwithin the individual vesicular agents or may be incorporated within amatrix formed by the biodegradable material of the microbeads butoutside the individual vesicular agents. Further embodiments of thisdisclosure are directed to methods for preparing the drug-loadedmicrobead compositions, to embolization compositions prepared from thedrug-loaded microbead compositions, to methods for preparing theembolization compositions, and to methods for treating a disease usingthe embolization compositions. The drug-loaded microbead compositionsinclude microbeads into which a therapeutic agent has been pre-loaded.When included in an embolization composition and provided to a subjectas part of an embolization therapy, the embolization compositions mayprovide a continuous release of the drug into the target treatment sitefor at least 7 days, at least 14 days, at least 21 days, or at least 30days after implantation.

According to embodiments, drug-loaded microbead compositions includeplurality of individual microbeads of a biodegradable material, aplurality of individual vesicular agents, a first therapeutic agent, anda second therapeutic agent different from the first therapeutic agent.In some embodiments, the plurality of individual vesicular agents may becontained within the individual microbeads, such as within pores of theindividual microbeads. In some embodiments, the plurality of individualvesicular agents may be associated with the biodegradable material ofthe individual microbeads through ionic or non-covalent interaction,either entirely encompassed with a matrix of the biodegradable materialor attached to the biodegradable material such as on an outer surface ofthe individual microbeads, for example. The vesicular agents include atleast one lipid bilayer surrounding a vesicular core. The firsttherapeutic agent is associated with the individual vesicular agents.For example, the first therapeutic agent may be contained within theindividual vesicular agents or may be associated with the individualvesicular agents by ionic or non-covalent interaction. The secondtherapeutic agent is different from the first therapeutic agent and iscontained within the individual microbeads or associated with theindividual microbeads through ionic or non-covalent interaction.

With respect to the vesicular agents and the therapeutic agents, theterm “associated with” includes non-covalent interactions such as ionicbonds, van der Waals interactions, hydrogen bonding, pi-pi stacking,dipole-dipole interactions, dipole-quadrupole interactions,quadrupole-quadrupole interactions, multipole-multipole interactions, orcombinations thereof. Ionic interactions or ionic bonding include a typeof chemical bonding that involves the electrostatic attraction betweenoppositely charged ions such as anions or cations. In some examples, atherapeutic agent may include atoms or groups capable of forming ions ormay have one or more natural dipoles. In such examples, the ions ordipoles may have particular affinity toward an oppositely charged ion ordipole naturally present in the vesicular agent or the biodegradablematerial. This affinity, in turn, results in a reversible chemicalattraction or combination between the therapeutic agent and thevesicular agent or the biodegradable material.

The individual microbeads of the drug-loaded microbead compositions aremicrobeads of a biodegradable material. The biodegradable material orthe individual microbeads forms a bead matrix that may include pores andmay have various shapes or sizes. In some embodiments, the microbeadsare spherical or ovoid. In some embodiments, the biodegradable materialis a biodegradable polymer or a lipid. Examples of biodegradablepolymers include, without limitation, poly(lactide-co-glycolide) (PLGA),polylactide (PLA), polyglycolide (PGA), polycaprolactone (PCL),polyhydroxyalkanoates, poly-R-3-hydroxybutyrate (poly3HB),poly-R-3-hydroxy-butyrate-co-R-3-hydroxyvalerate (poly(3HB-co-3HV)),poly-R-3-hydroxybutyrate-co-4-hydroxybutyrate (poly(3HB-co-4HB)),poly-R-3-hydroxyoctanoate-co-R-3-hydroxy-hexanoate (poly(3HO-co-3HH)),poly-3-hydroxypropionate (poly(3HP)), poly-4-hydroxy-butyrate(poly(4HB)), poly-5-hydroxybutyrate (poly(5HB)), poly-6-hydroxy-butyrate(poly(6HB)), poly(propylene fumarate), poly(butylene succinate),poly(p-dioxanone, polyacetals, poly(ortho esters), polycarbonates,chitosan, hydroxybutyric acids, polyanhydrides and polyesters,polyphosphazenes, polyphosphoesters, lipodisq, celluloses, modifiedcelluloses, proteins and poly(amino acids), polyethers, and co-polymersof the foregoing. Examples of lipids include, without limitation,tricaprin, trilaurin, trimyristin, tripalmitin, tristearin, hydrogenatedcoco-glycerides, glyceryl monostearate, glyceryl behenate, glycerylpalmitostearate, glyceryl caprate, cetyl palmitate, stearic acid,palmitic acid, decanoic acid, behenic acid, beeswax, carnauba wax, cacaobutter, and combinations thereof. In example embodiments, thedrug-loaded microbead compositions include individual microbeads arebeads of a biodegradable cellulose or a biodegradable modified cellulosesuch as cellulose acetate butyrate.

The biodegradable polymer materials of the drug-loaded microbeadcomposition may include derivatives of any of the foregoing materials,or may include combinations of any of the foregoing materials or theirderivatives. For example, the drug-loaded microbead compositions mayinclude a combination of multiple biodegradable polymer materials, inwhich each individual microbead is made of a single type of polymer, andthe drug-loaded microbead composition includes microbeads of multiplepolymer types. Alternatively, the drug-loaded microbead compositions mayinclude a combination of multiple biodegradable polymer materials, inwhich individual microbeads of the composition are composed of multipletypes of polymer.

In various embodiments, the biodegradable material of the drug-loadedmicrobead composition may include a solid lipid such as tricaprin,trilaurin, trimyristin, tripalmitin, tristearin, hydrogenatedcoco-glycerides, glyceryl monostearate, glyceryl behenate, glycerylpalmitostearate, glyceryl caprate, cetyl palmitate, stearic acid,palmitic acid, decanoic acid, behenic acid, beeswax, carnauba wax, andcacao butter. The biodegradable materials of the drug-loaded microbeadcomposition may include derivatives of any of the foregoing solidlipids, or may include combinations of any of the foregoing solid lipidsor their derivatives. For example, the drug-loaded microbeadcompositions may include a combination of multiple solid lipids, inwhich each individual microbead is made of a single type of solid lipid,and the drug-loaded microbead composition includes microbeads ofmultiple solid lipids. Alternatively, the drug-loaded microbeadcompositions may include a combination of multiple solid lipids, inwhich individual microbeads of the composition are composed of multipletypes of solid lipids.

In the drug-loaded microbead compositions according to embodiments,examples of vesicular agents include liposomes and ethosomes. Liposomesand ethosomes are particles having at least one amphipathic, sphericalor nearly spherical bilayer formed by van der Waals interactions betweena plurality of hydrophobic moieties each capped by a polar head groupand arranged in an alternating manner such that a polar head group ofone hydrophobic moiety projects outwardly to an external aqueousenvironment, while an adjacent hydrophobic moiety projects its polarhead group inwardly. Liposomes can be classified according to theirlamellarity (uni- and multilamellar vesicles), size (small,intermediate, or large), and charge (anionic, cationic and neutral) ofthe polar head groups. Liposome particles typically have diameters fromabout 0.025 μm to about 2.5 μm, in which the hydrophobic moieties arelinear or lightly branched saturated hydrocarbons. Ethosomes aredistinguished from liposomes in that the vesicular core of ethosomesincludes aqueous ethanol solution. In embodiments, a therapeutic agentmay be dissolved within the aqueous ethanol solution of ethosomes.

The vesicular agents such as liposomes or ethosomes, for example,include at least one lipid bilayer. The lipid bilayer may include aphospholipid. Examples of phospholipids include, without limitation,poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), soybeanphosphatidylcholine (SW), hydrogenated soybean phosphatidylcholine(HSPC), egg sphingomyelin (ESM), egg phosphatidylcholine (EPC),dimyristoyl phosphatidylcholine (DMPC), dipalmitoyl phosphatidylcholine(DPPC), dioleoyl phosphatidylcholine (DOPC), distearoylphosphatidylcholine (DSPC), dimyristoyl phosphatidylglycerol (DMPG),dipalmitoyl phosphatidylglycerol (DPPG), dioleoyl phosphatidylglycerol(DOPG), distearoyl phosphatidylglycerol (DSPC), dimyristoylphosphatidylethanolamine (DMPE), dipalmitoyl phosphatidylethanolamine(DPPE), dioleoyl phosphatidylethanolamine (DOPE), dimyristoylphosphatidylserine (LAMPS), dipalmitoyl phosphatidylserine (DPPS),dioleoyl phosphatidylserine (LOPS),1,2-dioleoyl-3-trimethylammonium-propane (DOTAP),1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC),1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC),1,2-dimyristoyl-sn-glycero-3-phosphate (sodium salt) (DMPA.Na),1,2-dipalmitoyl-sn-glycero-3-phosphate (sodium salt) (DPPA.Na),1,2-dioleoyl-sn-glycero-3-phosphate (sodium salt) (DOPA.Na),1,2-dimyristoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (sodium salt)(DMPG.Na), 1,2-dipalmitoyl-sn-glycero-3-phospho-(1′-rac-glycerol)(sodium salt) (DPPG.Na),1,2-dioleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (sodium salt)(DOPG.Na), 1,2-dimyristoyl-sn-glycero-3-phospho-L-serine (sodium salt)(DMPS.Na), 1,2-dipalmitoyl-sn-glycero-3-phospho-L-serine (sodium salt)(DPPS.Na), 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (sodium salt)(DOPS.Na), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(glutaryl)(sodium salt) (DOPE-glutaryl (Na)₂),1′,3′-bis[1,2-dimyristoyl-sn-glycero-3-phospho]-sn-glycerol (ammoniumsalt) (Tetramyristoyl Cardiolipin1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol)-2000] (ammonium salt) (DSPE-mPEG-2000, Na),1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethyleneglycol)-5000] (ammonium salt) (DSPE-mPEG-5000.Na),1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[maleimide(polyethyleneglycol)-2000] (ammonium salt) (DSPE-Maleimide PEG-2000.Na),1,2-dioleoyl-3-trimethylammonium-propane (chloride salt) (DOTAP.Cl),1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC), and mixtures thereof,and salts thereof. In example embodiments, the lipid bilayer of thevesicle agents such as liposomes or ethosomes may be a combination oflipids. In a specific embodiment, the lipid bilayer of the vesicleagents such as liposomes or ethosomes may be a combination ofpoly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), dioleoylphosphatidylserine (DOPS), dioleoyl phosphatidylethanolamine (DOPE), andcholesterol.

The individual microbeads of the drug-loaded microbead compositionsaccording to embodiments include at least two therapeutic agents, suchas the first therapeutic agent and the second therapeutic agent. Thefirst therapeutic agent is associated with or carried within theindividual vesicular agents. For example, the first therapeutic agentmay be contained within the individual vesicular agents or may beassociated with the individual vesicular agents by ionic or non-covalentinteraction. The first therapeutic agent may be within the lipid bilayerof the vesicular agents, within the vesicular core of the vesicularagents, or bound to the outer surface of the lipid bilayer and,therefore, not encapsulated by the vesicular agents.

The second therapeutic agent is different from the first therapeuticagent. The second therapeutic agent is contained within the individualmicrobeads or associated with the individual microbeads through ionic ornon-covalent interaction. In some embodiments, the second therapeuticagent is contained within the individual vesicular agents of theindividual microbeads or associated with individual vesicular agentsthrough ionic or non-covalent interaction. For example, in someembodiments, the second therapeutic agent may be within the lipidbilayer of the vesicular agents, within the vesicular core of thevesicular agents, or bound to the outer surface of the lipid bilayerand, therefore, not encapsulated by the vesicular agents. In someembodiments, the second therapeutic agent is not contained within theindividual vesicular agents and not ionically or non-covalentlyassociated with the individual vesicular agents but, rather, isassociated by ionic or non-covalent interaction with the biodegradablematerial of the individual microbeads. In such embodiments, theindividual microbeads may contain vesicular agents loaded with the firsttherapeutic agent and molecules of the second therapeutic agent insideor attached to the matrix of biodegradable material that forms theindividual microbeads but not inside the vesicular agents.

In some embodiments of the drug-loaded microbead composition the firsttherapeutic agent is hydrophobic and the second therapeutic agent ishydrophilic. In example embodiments, the first therapeutic agent may beencapsulated within the vesicular core of the individual vesicularagents and the second therapeutic agent may be contained within thelipid bilayer of the individual vesicular agents. In further exampleembodiments, the first therapeutic agent may be encapsulated within thevesicular core of the individual vesicular agents and the secondtherapeutic agent is not contained within the individual vesicularagents and is associated by ionic or non-covalent interaction with thebiodegradable material of the individual microbeads.

The drug-loaded microbead compositions include at least two therapeuticagents. In embodiments, the first therapeutic agent and the secondtherapeutic agent are independently chosen from doxorubicin,bevacizumab, bortezomib, imatinib, seliciclib, ceritinib, everolimus,paclitaxel, sorafenib, irinotecan, idarubicin, cisplatin,pharmaceutically acceptable salts and derivatives thereof, andcombinations thereof, provided the first therapeutic agent is differentfrom the second therapeutic agent. In some embodiments, the therapeuticagent may be a hydrophilic therapeutic agent or a therapeutic agent thateither is water soluble or has at least some solubility in an aqueoussolution. In some embodiments, the therapeutic agent may be achemotherapeutic agent having at least some efficacy for treating adisease such as cancer. In some embodiments, the therapeutic agent maybe a chemotherapeutic agent having at least some efficacy for treating acancer such as hepatocellular carcinoma, liver cancer, prostate cancer,or breast cancer. The therapeutic agent may have one or more chemicalmoieties or atomic centers having a positive or negative charge oraffinity. Examples of specific therapeutic agents include, withoutlimitation, doxorubicin, sorafenib, vandetanib, nivolumab, ipilimumab,regorafenib, irinotecan, epirubicin, pirarubicin, 5-fluorouracil,cisplatin, floxuridine, mitomycin C, derivatives of any of theforegoing, prodrugs of any of the foregoing, therapeutically acceptablesalts or crystalline forms of any of the foregoing, or combinations ofany of the foregoing. Further examples of suitable therapeutic agentsinclude, without limitation, pirarubicin, mitoxantrone, tepotecan,paclitaxel, carboplatin, pemetrexed, penistatin, pertuzumab,trastuzumab, and docetaxel. Still further examples of suitabletherapeutic agents include, without limitation, doxorubicin,bevacizumab, bortezomib, imatinib, seliciclib, ceritinib, everolimus,paclitaxel, sorafenib, irinotecan, idarubicin, cisplatin, andcombinations of these drugs.

In an example embodiment of a drug-loaded microbead composition, theindividual vesicular agents include liposomes; the first therapeuticagent is contained within the lipid bilayer of the individual vesicularagents; the second therapeutic agent is not contained within theindividual vesicular agents and is associated by ionic or non-covalentinteraction with the biodegradable material of the individualmicrobeads; and the biodegradable material is a cellulose. In suchembodiments, the liposomes may include a lipid bilayer formed from amixture of poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC),dioleoyl phosphatidylserine (DOPS), dioleoyl phosphatidylethanolamine(DOPE), and cholesterol, for example.

In a further example embodiment of a drug-loaded microbead composition,the individual vesicular agents include liposomes; the first therapeuticagent is contained within the lipid bilayer of the individual vesicularagents; the second therapeutic agent is encapsulated within thevesicular core of the individual vesicular agents; the first therapeuticagent is sorafenib; the second therapeutic agent is doxorubicin; and thebiodegradable material is a cellulose or modified cellulose such asethyl cellulose or cellulose acetate butyrate. In such embodiments, theliposomes may include a lipid bilayer formed from a mixture ofpoly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), dioleoylphosphatidylserine (DOPS), dioleoyl phosphatidylethanolamine (DOPE), andcholesterol, for example.

In a further example embodiment of a drug-loaded microbead composition,the individual vesicular agents include ethosomes; the first therapeuticagent is encapsulated within the vesicular core of the individualvesicular agents; the second therapeutic agent is not contained withinthe individual vesicular agents and is associated by ionic ornon-covalent interaction with the biodegradable material; the firsttherapeutic agent is sorafenib; the second therapeutic agent isdoxorubicin; and the biodegradable material is a cellulose or modifiedcellulose such as ethyl cellulose or cellulose acetate butyrate. In suchembodiments, the liposomes may include a lipid bilayer formed from amixture of poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC),dioleoyl phosphatidylserine (DOPS), dioleoyl phosphatidylethanolamine(DOPE), and cholesterol, for example.

In the individual microbeads of the drug-loaded microbead composition,the therapeutic agent or a complex of the therapeutic agent may beencapsulated by or embedded within the matrix formed by thebiodegradable material. Except when described otherwise, the terms“encapsulated” and “embedded” broadly include embodiments for which thebiodegradable material or some portion thereof generally surrounds thetherapeutic agent and/or the vesicular agents including a therapeuticagent. For example, “embedded” may include in some embodiments abiodegradable material shell that encapsulates a core that holds thetherapeutic agent and/or vesicular agents. In other embodiments,“embedded” may include a structure in which the therapeutic agent or thecomplex is physically disposed within a matrix, network, or porestructure of a biodegradable material that may or may not have a corewithin an outer shell. The therapeutic agent itself may not bechemically bonded to the biodegradable material at all or may not bechemically bonded directly to the polymer backbone of the biodegradablematerial when the biodegradable material is a biodegradable polymermaterial. The drug-loaded microbead composition may have a water contentof less than 1% by weight, based on the total weight of the drug-loadedmicrobead composition.

In some embodiments, the therapeutic agent of the drug-loaded microbeadcomposition may be embedded within the microbeads of the biodegradablematerial but not chemically bonded to the biodegradable material. Insome embodiments, the therapeutic agent of the drug-loaded microbeadcomposition may be embedded within the microbeads of the biodegradablematerial and not chemically bonded directly to a polymer backbone of thebiodegradable material, yet may be chemically bonded to a functionalgroup of the biodegradable material. As used herein, “not chemicallybonded” refers to a lack of covalent chemical bonds between thetherapeutic agent and the biodegradable but does not preclude theexistence of noncovalent intermolecular interactions such as ionicinteractions or a van der Waals interaction between the therapeuticagent and the biodegradable material.

In some embodiments, the vesicular agents form complexes of thetherapeutic agent. In the complex, the therapeutic agent may bechemically bonded to the vesicular agents or the lipid bilayer thereofor may be associated with the vesicular agents or the lipid bilayerthereof by a non-covalent means such as encapsulation or a van der Waalsinteraction. The complex may be embedded within the biodegradablematerial. When the complex is embedded within the microbeads, thevesicular agents may be chemically bonded to the biodegradable materialwhile the therapeutic agent is not chemically bonded to thebiodegradable material. Without intent to be bound by theory, it isbelieved that when the therapeutic agent is bonded or associated withthe vesicular agents but is not chemically bonded to the biodegradablematerial, the microbeads of the drug-loaded microbead composition areless susceptible to shrinking as a result of replacing water moleculeswith drug molecules during drug loading. Accordingly, the final sizedistribution of the drug-loaded microbeads can be controlled morereadily by selecting appropriate microbead sizes before the therapeuticagent is loaded.

The drug-loaded microbead compositions according to embodiments may havea very low water content such as less than 1%, or less than 0.1%, orless than 0.05% (500 ppm), or less than 0.02% (200 ppm), or less than0.01% (100 ppm), or less than 0.005 (50 ppm), or less than 0.002% (20ppm), or less than 0.001% (10 ppm) by weight water, based on the totalweight of the microbeads in the drug-loaded microbead composition.Without intent to be bound by theory, it is believed that a very lowwater content of the drug-loaded microbead composition increases theshelf-life and long-term stability of the drug-loaded microbeadcomposition. Further, it is believed that water contents significantlygreater than 1% by weight (such as 2%, 3%, 5%, or 10%, for example)based on the total weight of the drug-loaded microbead composition, maylead to decomposition or hydrolysis of the therapeutic agent,instability or breaking apart of the water-swellable polymer, or acombination of these, within a few days or even a few hours, such thatthe composition cannot be used for embolization procedures, even if thedrug-loaded microbead composition is rehydrated. It is believed that theshelf-life and long-term stability of compositions having water contentssignificantly greater than 1% by weight are not sufficiently long toensure viability of the therapeutic agent over the time period frommanufacture of the drug-loaded microbead composition to use of thecomposition in an embolization procedure. It is believed that selectionof the water-swellable polymer material may correlate with the abilityfor water to be removed from the drug-loaded microbeads bylyophilization or other drying technique or combination of dryingtechniques in an amount sufficient to prevent decomposition of thetherapeutic agent.

A very low water content of the drug-loaded microbead composition, aspreviously described may be attained by drying techniques that will bedescribed in greater detail subsequently, with respect to methods forpreparing the drug-loaded microbead compositions. In this regard, thedrug-loaded microbead compositions may be dry or nearly dehydratedcompositions of the microbeads containing the embedded therapeutic agentor the embedded complex of the therapeutic agent and the vesicularagents. The drug-loaded microbead compositions may have a powder-likeconsistency. Accordingly, the drug-loaded microbead compositions may bemade suitable for injection into a subject being treated by rehydratingthe microbeads of the drug-loaded microbead compositions to form anembolization composition, as will be described in greater detailsubsequently. Regardless, the drug-loaded microbead compositions may beprovided in such a form that a physician needs to add only an aqueoussolution such as water or physiologically buffered saline solution tothe drug-loaded microbead composition to prepare the composition for usein an embolization procedure.

The microbeads of the drug-loaded microbead composition may have anyshape common to microparticles formed from a hydrogel typewater-swellable polymer material. For example, the microparticles may bespherical or substantially spherical, or may have an ovoid shape withoval-shaped or elliptical cross-sections about a longitudinal axis andcircular cross-sections about an axis perpendicular to the longitudinalaxis.

The drug-loaded microbead composition may include an amount oftherapeutic agent per unit volume of microbeads in the composition thatis chosen to have a desired therapeutic effect or activity, based on theintended use for the drug-loaded microbead composition and theparticular therapeutic agent present in the individual microbeads.

The amount of therapeutic agent in the individual microbeads of thedrug-loaded microbead composition can be adjusted through particulartechniques involved during drug loading, such as loading time, loadingtemperature, or concentration of therapeutic agent in a loadingsolution, for example. The amount of therapeutic agent in the individualmicrobeads of the drug-loaded microbead composition can be adjusted alsothrough synthetic techniques involved for synthesizing the microbeadsthemselves, such as through adjusting polymer molecular weights, polymerdensity, or polymer porosity of the biodegradable material. For example,when doxorubicin is the therapeutic agent, the amount of drug loading inthe drug-loaded microbeads can be adjusted with respect to the number ofnegative charges in the polymer backbone of the water-swellable polymermaterial. Similarly, when sorafenib is the therapeutic agent, thesorafenib may be embedded within liposomes that are embedded within themicrobead structure.

In example embodiments, the individual drug-loaded microbeads of thedrug-loaded microbead composition include the biodegradable material,the first and second therapeutic agents, and water. The individualdrug-loaded microbeads of the drug-loaded microbead composition mayinclude from about 30% by weight to about 70% by weight, or from about35% by weight to about 65% by weight, or from about 40% to about 60% byweight, or about 45% by weight to about 55% by weight, or about 50% toabout 70% by weight biodegradable material, based on the total weight ofthe individual drug-loaded microbead. In example embodiments, theindividual drug-loaded microbeads of the drug-loaded microbeadcomposition may include from about 1% by weight to about 25% by weight,or from about 1% by weight to about 20% by weight, or from about 1% byweight to about 15% by weight, or from about 2% by weight to about 25%by weight, or from about 5% by weight to about 25% by weight, or fromabout 10% by weight to about 25% by weight therapeutic agent, based onthe total weight of the individual drug-loaded microbead. In exampleembodiments, the individual drug-loaded microbeads of the drug-loadedmicrobead composition according to embodiments may have a very low watercontent such as less than 1% by weight, or less than 0.5% by weight, orless than 0.1% by weight, or less than 0.05% (500 ppm) by weight, orless than 0.02% (200 ppm) by weight, or less than 0.01% (100 ppm) byweight, or less than 0.005 (50 ppm) by weight, or less than 0.002% (20ppm) by weight, or less than 0.001% (10 ppm) by weight water, based onthe total weight of the individual microbeads in the drug-loadedmicrobead composition.

Having described drug-loaded microbead compositions according to variousembodiments, methods for preparing the drug-loaded microbeadcompositions will now be described.

Example methods for preparing drug-loaded microbead compositions includepreparing vesicular agents, adding the vesicular agents to an aqueoussolution, preparing an organic solution containing a precursor to abiodegradable material, combining the aqueous solution and the organicsolution, agitating the mixture to form microbeads, drying themicrobeads, and recovering the microbeads. The methods may furtherinclude washing, rinsing, or filtering the microbeads.

The foregoing methods for preparing drug-loaded microbead compositions,regardless of whether the drug-loaded microbeads are prepared includingthe intermediate step of forming particles of the complexes oftherapeutic agent and vesicular agents or without the intermediate step,further include removing water from the drug-loaded microbeads to form adrug-loaded microbead composition having a water content of less than 1%by weight, based on the total weight of the drug-loaded microbeadcomposition. In some embodiments, the removal of the water may includelyophilization or freeze drying, optionally followed by an additionaldrying step involving temperature variation (heating or cooling),flowing air, vacuum, or a combination of these. The drug-loadedmicrobeads may have an average synthesized volume upon initial recoveryand an average final volume after the removing of water. The averagefinal volume may range from about 10% to about 75% of the averagesynthesized volume such as, for example, about 10%, about 20%, about30%, about 40%, about 50%, about 60%, about 70%, or about 75% of theaverage synthesized volume.

Lyophilization or freeze drying of the drug-loaded microbeads may beconducted according to any procedure commonly used for drying ofparticulate substances. For example, the drug-loaded microbeads may beplaced in partially stoppered glass vials, which then are placed on acooled, temperature-controlled shelf within a freeze dryer. The shelftemperature is reduced, and the sample is frozen to a uniform, definedtemperature. After complete freezing, the pressure in the dryer may bedecreased to a defined pressure to initiate primary drying. During theprimary drying, water vapor is progressively removed from the frozenmass by sublimation while the shelf temperature is controlled at aconstant, low temperature. Secondary drying may be initiated byincreasing the shelf temperature and reducing the chamber pressurefurther so that water absorbed to the semi-dried mass can be removeduntil the residual water content decreases to the desired level.

Lyophilization or freeze drying of the drug-loaded microbeads may alsobe conducted by atmospheric pressure freeze drying by rapidlycirculating very dry air over the frozen drug-loaded microbeads. Thecirculating dry gas provides improved heat and mass transfer from thefrozen microbeads. Atmospheric spray drying processes may facilitate theformation of small-diameter microbeads as a free-flowing powder, whileavoiding formation of a dried cake. The free-flowing powder, in turn,may facilitate rehydration of the drug-loaded microbead composition whenan embolization composition is prepared.

The drug-loaded microbeads of the drug-loaded microbead compositionspreviously described, or prepared according to the foregoing methods,may exhibit physical or mechanical properties beneficial to their use,storage, transport, or subsequent rehydration to form an embolizationcomposition.

The drug-loaded microbeads of the drug-loaded microbead compositionspreviously described, or prepared according to the foregoing methods,may be provided to a physician with instructions for rehydrating thedrug-loaded microbead composition to form an embolization compositionthat is ready for use in an embolization treatment. Accordingly,embodiments of this disclosure include embolization compositions andmethods for preparing the embolization compositions.

An embolization composition may include a drug-loaded microbeadcomposition according to any embodiment of this disclosure or adrug-loaded microbead composition prepared according to an embodiedmethod of this disclosure. The embolization composition may furtherinclude amount of an aqueous solution sufficient to cause thedrug-loaded microbeads of the drug-loaded microbead composition toreconstitute to a form suitable for administration to a patient. Inparticular, the amount of aqueous solution may cause the drug-loadedmicrobeads, after hydration, to have a water content of from about 50%by weight to about 99% by weight, from about 60% by weight to about 95%by weight, from about 70% by weight to about 95% by weight, from about80% by weight to about 95% by weight, from about 90% by weight to about95% by weight, or from about 80% by weight to about 99% by weight, orfrom about 90% by weight to about 99% by weight, or from about 95% byweight to about 99% by weight, based on the total weight of thedrug-loaded microbeads. The aqueous solution may be any pharmaceuticallyacceptable solution such as, for, example, a physiologically bufferedsaline solution.

In example embodiments, the hydrated individual drug-loaded microbeadsof the embolization composition include the biodegradable material, thetherapeutic agent, the vesicular agents, and water. The hydratedindividual drug-loaded microbeads of the embolization composition mayinclude from about 3% by weight to about 10% by weight, from about 3% byweight to about 7% by weight, or from about 3.5% by weight to about 6.5%by weight, or from about 4% to about 6% by weight, or about 4.5% byweight to about 5.5% by weight, or about 5% to about 7% by weightbiodegradable material, based on the total weight of the hydratedindividual drug-loaded microbead. In example embodiments, the hydratedindividual drug-loaded microbeads of the embolization composition mayinclude from about 0.1% by weight to about 2.5% by weight, or from about0.1% by weight to about 2% by weight, or from about 0.1% by weight toabout 1.5% by weight, or from about 0.2% by weight to about 2.5% byweight, or from about 0.5% by weight to about 2.5% by weight, or fromabout 1% by weight to about 2.5% by weight therapeutic agent, based onthe total weight of the hydrated individual drug-loaded microbead. Inexample embodiments, the hydrated individual drug-loaded microbeads ofthe embolization composition according to embodiments may have a watercontent of from about 50% by weight to about 99% by weight, from about50% by weight to about 97% by weight, from about 50% by weight to about96% by weight, from about 60% by weight to about 95% by weight, fromabout 70% by weight to about 95% by weight, from about 80% by weight toabout 95% by weight, from about 90% by weight to about 95% by weight, orfrom about 80% by weight to about 99% by weight, or from about 90% byweight to about 99% by weight, or from about 95% by weight to about 99%by weight, based on the total weight of the individual drug-loadedmicrobeads.

The microbeads of the embolization composition may have averagediameters from about 5 μm to about 1200 μm, as measured by dynamic lightscattering (DLS). For non-spherical microbeads, the diameter of anindividual particle is taken to be the widest measurement attainablefrom a first point on the surface of the microbead, through the centerof mass of the microbead, to a second point on the surface of themicrobead opposite the first point. The specific size distribution ofthe microparticles may be chosen or tailored to suit the particulartreatment for which the embolization composition is intended to be used.The average diameters of the microbeads in the embolization compositionmay be chosen by any suitable method of size selection before or afterthe therapeutic agent is loaded into the microbeads. In someembodiments, the swollen microbeads of the embolization composition mayhave average diameters such as, for example, from about 40 μm to about800 μm, from about 40 μm to about 100 μm, from about 40 μm to about 75μm, from about 75 μm to about 100 μm, from about 100 μm to about 200 μm,from about 200 μm to about 300 μm, from about 300 μm to about 400 μm,from about 400 μm to about 500 μm, from about 600 μm to about 700 μm, orany subset of any of the foregoing ranges. In some embodiments, theswollen microbeads may have average diameters with a narrow sizedistribution such as 40 μm±20 μm, or 40 μm±10 μm, or 40 μm±5 μm, or 40μm±1 μm, for example.

Methods for preparing an embolization composition may include adding adrug-loaded microbead composition according to any embodiment of thisdisclosure, or a drug-loaded microbead composition prepared according toan embodied method of this disclosure, to an amount of an aqueoussolution sufficient to cause the drug-loaded microbeads of thedrug-loaded microbead composition to swell, whereby the drug-loadedmicrobeads after swelling comprise a water content of from 50% to 99% byweight, from 50% to 90% by weight, from 50% to 75% by weight, from 60%to 99% by weight, from 75% to 99% by weight, from 75% to 95% by weight,from 75% to 90% by weight, or from 85% to 99% by weight, based on thetotal weight of the drug-loaded microbeads, to form an injection-readysolution. The aqueous solution may be any pharmaceutically acceptablesolution such as, for, example, a physiologically buffered salinesolution. The methods may further include loading the injection-readysolution into an injection device such as a syringe, for example. Themethods may further include allowing the drug-loaded microbeads to swellfor a rehydration time of from about 5 minutes to about 60 minutesbefore loading the injection-ready solution into the injection device.In some examples, the embolization composition may contain from about 25mg therapeutic agent per milliliter of microbeads to about 150 mgtherapeutic agent per milliliter of microbeads.

The embolization compositions prepared from the drug-loaded microbeadcompositions according to embodiments of this disclosure may beincorporated into an embolization treatment or therapy intended to treata disease such as a cancer. In some embodiments, the therapeutic agentmay be a chemotherapeutic agent having at least some efficacy fortreating a cancer such as hepatocellular carcinoma, liver cancer,prostate cancer, or breast cancer. Accordingly, embodiments of thisdisclosure include methods for treating a disease. Methods for treatinga disease may include providing intravenously, to a subject in need ofembolization therapy, an embolization composition according to anyembodiment previously described herein, prepared from a drug-loadedmicrobead composition according to any embodiment previously describedherein.

In the methods for treating a disease, after the embolizationcomposition is provided intravenously, at least a portion of thedrug-loaded microbeads flow through vasculature of the subject to anembolization site and restrict blood flow through the embolization site.Thereafter, the drug-loaded microbeads may release to tissue at theembolization site over the course of a release period at least 90% byweight of the therapeutic agent, based on an initial amount oftherapeutic agent embedded within the microbeads before the embolizationcomposition is provided. In some embodiments, the release to the tissueat the embolization site may include an initial burst of drug release,during which at least 10% by weight of the therapeutic agent initiallypresent in the drug-loaded microbeads is released into surroundingtissues at the embolization site. The initial burst may occur within 1minute, 5 minutes, 10 minutes, 30 minutes, 60 minutes, 120 minutes, 180minutes, 240 minutes, 300 minutes, 360 minutes, 12 hours, 18 hours, or24 hours after the drug-loaded microbeads reach the embolization site.In some embodiments, the drug-loaded microbeads provide a sustainedrelease of the therapeutic agent to the tissue over the release period.The release period may be an extended release period such as at least 5days, at least 10 days, at least 14 days, at least 28 days, or at least42 days. The end of release period is determined from the time when thedrug-loaded microbeads are either completely decomposed or stop elutingtherapeutic agent to the tissue.

Without intent to be bound by theory, it is believed that the releasemechanism of the therapeutic agent from the drug-loaded microbeads ofthe embolization composition may be based on a dual mechanism includingan ion exchange process and an enzymatic process. For example, it isbelieved that in an early stage of delivery of therapeutic agent to theembolization site, when plenty of water is present, the therapeuticagent may be released from the drug-loaded microbeads by ion exchangeprocess. In a later stage, when water is lacking due to the vessel'sbeing embolized, the drug-loaded microbeads may be degraded by anenzymatic process such as by lysozymes, for example, to release thetherapeutic agent from the microbeads. It is believed that in the lateststage, such as by day 21 following implantation, most of thebiodegradable material has been resorbed through tissue surrounding theembolization site. Because at this point the water-swellable material nolonger has a matrix encapsulating the therapeutic agent or the complexof the therapeutic agent and the vesicular agents, the therapeutic agentcan be completely eluted into the embolization site. Thereby, it isbelieved that the embolization compositions according to embodiments ofthis disclosure may increase tumor response and free survival rates.Additionally, the embolization compositions according to embodiments ofthis disclosure provide an economic value such as reducing the physicianprocedure time by eliminating a need to request a pharmacist ortechnician to add therapeutic agent to unloaded microbeads.

Further embodiments of this disclosure are directed to drug-loadedbiodegradable microbead compositions. The drug-loaded biodegradablemicrobead compositions may include microbeads of a biodegradablematerial and at least one therapeutic agent embedded in a matrix of thebiodegradable material, encapsulated by a matrix of the biodegradablematerial, disposed within a porous structure formed by the matrix of thebiodegradable material, or ionically or non-covalently associated with amatrix of the biodegradable material. In such embodiments of drug-loadedbiodegradable microbead compositions, the microbeads of thebiodegradable material may contain, but need not necessarily contain,vesicular agents such as liposomes or ethosomes.

In the drug-loaded biodegradable microbead compositions, thebiodegradable material is a biodegradable polymer or a lipid. Examplesof biodegradable polymers include, without limitation,poly(lactide-co-glycolide) (PLGA), polylactide (PLA), polyglycolide(PGA), polycaprolactone (PCL), polyhydroxyalkanoates,poly-R-3-hydroxybutyrate (poly3HB),poly-R-3-hydroxybutyrate-co-R-3-hydroxyvalerate (poly(3HB-co-3HV)),poly-R-3-hydroxybutyrate-co-4-hydroxybutyrate (poly(3HB-co-4HB)),poly-R-3-hydroxyoctanoate-co-R-3-hydroxyhexanoate (poly(3HO-co-3HH)),poly-3-hydroxypropionate (poly(3HP)), poly-4-hydroxybutyrate(poly(4HB)), poly-5-hydroxybutyrate (poly(5HB)), poly-6-hydroxybutyrate(poly(6HB)), poly(propylene fumarate), poly(butylene succinate),poly(p-dioxanone, polyacetals, poly(ortho esters), polycarbonates,chitosan, hydroxybutyric acids, polyanhydrides and polyesters,polyphosphazenes, polyphosphoesters, lipodisq, celluloses, modifiedcelluloses, proteins and poly(amino acids), polyethers, and co-polymersof the foregoing. Examples of lipids include, without limitation,tricaprin, trilaurin, trimyristin, tripalmitin, tristearin, hydrogenatedcoco-glycerides, glyceryl monostearate, glyceryl behenate, glycerylpalmitostearate, glyceryl caprate, cetyl palmitate, stearic acid,palmitic acid, decanoic acid, behenic acid, beeswax, carnauba wax, cacaobutter, and combinations thereof. In example embodiments, thedrug-loaded microbead compositions include individual microbeads arebeads of a biodegradable cellulose or a biodegradable modified cellulosesuch as ethyl cellulose or cellulose acetate butyrate. The biodegradablepolymer materials of the drug-loaded biodegradable microbead compositionmay include derivatives of any of the foregoing materials, or mayinclude combinations of any of the foregoing materials or theirderivatives. For example, the drug-loaded microbead compositions mayinclude a combination of multiple biodegradable polymer materials, inwhich each individual microbead is made of a single type of polymer, andthe drug-loaded biodegradable microbead composition includes microbeadsof multiple polymer types. Alternatively, the drug-loaded biodegradablemicrobead compositions may include a combination of multiplebiodegradable polymer materials, in which individual microbeads of thecomposition are composed of multiple types of polymer.

In various embodiments, the biodegradable material of the drug-loadedbiodegradable microbead composition may include a solid lipid such astricaprin, trilaurin, trimyristin, tripalmitin, tristearin, hydrogenatedcoco-glycerides, glyceryl monostearate, glyceryl behenate, glycerylpalmitostearate, glyceryl caprate, cetyl palmitate, stearic acid,palmitic acid, decanoic acid, behenic acid, beeswax, carnauba wax, andcacao butter. The biodegradable materials of the drug-loadedbiodegradable microbead composition may include derivatives of any ofthe foregoing solid lipids, or may include combinations of any of theforegoing solid lipids or their derivatives. For example, thedrug-loaded biodegradable microbead compositions may include acombination of multiple solid lipids, in which each individual microbeadis made of a single type of solid lipid, and the drug-loadedbiodegradable microbead composition includes microbeads of multiplesolid lipids. Alternatively, the drug-loaded biodegradable microbeadcompositions may include a combination of multiple solid lipids, inwhich individual microbeads of the composition are composed of multipletypes of solid lipids.

In the drug-loaded biodegradable microbead compositions, the at leastone therapeutic agent may be chosen from doxorubicin, bevacizumab,bortezomib, imatinib, seliciclib, ceritinib, everolimus, paclitaxel,sorafenib, irinotecan, idarubicin, cisplatin, pharmaceuticallyacceptable salts and derivatives thereof, and combinations thereof.Further examples of specific therapeutic agents include, withoutlimitation, doxorubicin, sorafenib, vandetanib, nivolumab, ipilimumab,regorafenib, irinotecan, epirubicin, pirarubicin, 5-fluorouracil,cisplatin, floxuridine, mitomycin C, derivatives of any of theforegoing, prodrugs of any of the foregoing, therapeutically acceptablesalts or crystalline forms of any of the foregoing, or combinations ofany of the foregoing. Further examples of suitable therapeutic agentsinclude, without limitation, pirarubicin, mitoxantrone, tepotecan,paclitaxel, carboplatin, pemetrexed, penistatin, pertuzumab,trastuzumab, and docetaxel. Still further examples of suitabletherapeutic agents include, without limitation, doxorubicin,bevacizumab, bortezomib, imatinib, seliciclib, ceritinib, everolimus,paclitaxel, sorafenib, irinotecan, idarubicin, cisplatin, andcombinations of these drugs. In one specific embodiment of thedrug-loaded biodegradable microbead compositions, the at least onetherapeutic agent includes doxorubicin. In one specific embodiment ofthe drug-loaded biodegradable microbead compositions, the at least onetherapeutic agent includes sorafenib. In one specific embodiment of thedrug-loaded biodegradable microbead compositions, the at least onetherapeutic agent includes paclitaxel. In some embodiments, themicrobeads of the drug-loaded biodegradable microbead compositionsinclude combinations of two, three, four, or greater than fourtherapeutic agents. In one specific embodiment of the drug-loadedbiodegradable microbead compositions, the at least one therapeutic agentincludes a combination of doxorubicin and sorafenib. In someembodiments, the biodegradable material and the at least one therapeuticagent are both hydrophobic, thereby facilitating homogeneousdistribution of the therapeutic agent throughout the matrix of thebiodegradable material.

The microbeads of the drug-loaded biodegradable microbead compositionsmay be prepared by any suitable process including, but not limited to,emulsion and microfluidics. Emulsion methods include preparing anaqueous phase of water and, optionally, an emulsifier such as polyvinylalcohol; preparing an organic phase of a biodegradable material, ahydrophobic therapeutic agent, and a water-immiscible organic solvent;mixing the aqueous phase and the organic phase to form an emulsion;evaporating the solvents to dry the microbeads formed from the emulsion;recovering and further drying the microbeads; isolating desired sizeranges of beads by filtering or sieving; optionally further dryingand/or dehydrating and/or lyophilizing the microbeads; and sterilizingthe microbeads. The microbeads may be rehydrated or reconstituted byadding a fluid such as water or physiologically buffered saline.Microbeads formed in this manned may have sizes from 20 μm to 900 μm,for example, and can be sieved to narrow ranges. The ratio ofbiodegradable material to therapeutic agent by weight may be from 100:1to 0.01:1, or from 10:1 to 0.1:1, or any desired range between theseranges, for example. Microfluidics methods include similarly preparingaqueous and organic phases and, in a microfluidic apparatus, flowingsmall volumes of organic phase through the aqueous phase to producesmall droplets in the aqueous phase that may be collected as microbeadsat an exit of the apparatus.

Further embodiments of the present disclosure are directed to adrug-loaded microbead composition for use in a therapeutic embolizationprocedure. The drug-loaded microbead composition can be any of thedrug-loaded microbead compositions that are described herein, and can beused in the form of an embolization composition according to anyembodiment described herein. The therapeutic embolization procedure ispreferably one for treating tumors. The therapeutic embolizationprocedure is preferably one for treating hepatocellular carcinoma, livercancer, prostate cancer, or breast cancer.

EXAMPLES

The following examples are offered by way of illustration only. In viewof the foregoing description, the person having ordinary skill in theart will recognize that the following examples are not intended to limitthe scope of this disclosure or its many embodiments.

Example 1

A drug-loaded microbead composition of this Example includes polymermicrobeads and a complex comprising a dextran sulfate chemically bondedto doxorubicin. The complex is encapsulated within a core of themicrobead, defined within a shell of water-swellable polymer material.The microbeads have size ranges from 10 μm to 2000 μm. The drug-loadedmicrobead composition contains from about 25 mg doxorubicin permilliliter of microbeads to about 150 mg doxorubicin per milliliter ofmicrobeads.

To prepare the drug-loaded microbead composition, a dextran-doxorubicincomplex is prepared made by dissolving dextran sodium sulfate (molecularweight range 40 kDa to 500 kDa) and doxorubicin in an aqueous solution.Then the dextran-doxorubicin solution is added into a solvent mixturewith high speed stirring to generate doxorubicin-loaded dextranparticles. The doxorubicin-loaded particles are added into a hydrogelsolution of the polymer material and are encapsulated within themicrobead core by a water/oil process. In the water/oil process, thedoxorubicin-loaded dextran particles are present in an aqueous solvent,to which additional solvents are added to create a biphasic mixture. Thebiphasic mixture includes an aqueous phase, in which thedoxorubicin-loaded dextran particles are initially present with thehydrogel, and an oil phase. On rapid mixing of the biphasic mixture, thedoxorubicin-loaded dextran particles are incorporated into microbeads ofthe hydrogel polymer within the oil phase of the biphasic mixture.

After being recovered from the biphasic mixture, the drug-loadedmicrobeads contain a substantial amount of water and are in a swollenstate. To prepare the drug-loaded microbead composition for packaging,the microbeads are subjected to lyophilization to remove water. Thelyophilization process removes greater than 90% by weight, or greaterthan 95% by weight, or greater than 98%, or greater than 99% by weight,or greater than 99.9% by weight, or greater than 99.99% by weight of thetotal water present in the microbeads immediately after the drugloading. The lyophilized microbeads may be subjected to further dryingin air and/or vacuum. Thereby, the drug-loaded microbead compositioncontains less than 1%, or less than 0.1%, or less than 0.05% (500 ppm),or less than 0.02% (200 ppm), or less than 0.01% (100 ppm), or less than0.005 (50 ppm), or less than 0.002% (20 ppm), or less than 0.001% (10ppm) by weight water, based on the total weight of the drug-loadedmicrobead composition.

Example 2

A drug-loaded microbead composition of this Example also includespolymer microbeads and a complex comprising a dextran sulfate chemicallybonded to doxorubicin. The complex is encapsulated within a core of themicrobead, defined within a shell of water-swellable polymer material.The microbeads have size ranges from 10 μm to 2000 μm. The drug-loadedmicrobead composition contains from about 25 mg doxorubicin permilliliter of microbeads to about 150 mg doxorubicin per milliliter ofmicrobeads.

To prepare the drug-loaded microbead composition, a dextran-doxorubicincomplex is prepared made by dissolving dextran sodium sulfate (molecularweight range 40 kDa to 500 kDa) and doxorubicin in an aqueous solution.Unlike in Example 1, in which doxorubicin-loaded dextran particles aregenerated as an intermediate step, the dextran-doxorubicin solution iscombined directly with a hydrogel solution of the polymer material toform a synthesis mixture. After vigorous mixing, drug-loaded microbeadsform, in which the microbeads are embedded within an interpenetratingnetwork of water-swellable polymer material and dextran, and thedoxorubicin is complexed to the dextran without being chemically bondedto the water-swellable polymer material. The drug-loaded microbeads arerecovered from the synthesis mixture.

After being recovered from the synthesis mixture, the drug-loadedmicrobeads contain a substantial amount of water and are in a swollenstate. To prepare the drug-loaded microbead composition for packaging,the microbeads are subjected to lyophilization to remove water. Thelyophilization process removes greater than 90% by weight, or greaterthan 95% by weight, or greater than 98%, or greater than 99% by weight,or greater than 99.9% by weight, or greater than 99.99% by weight of thetotal water present in the microbeads immediately after the drugloading. The lyophilized microbeads may be subjected to further dryingin air and/or vacuum. Thereby, the drug-loaded microbead compositioncontains less than 1%, or less than 0.1%, or less than 0.05% (500 ppm),or less than 0.02% (200 ppm), or less than 0.01% (100 ppm), or less than0.005 (50 ppm), or less than 0.002% (20 ppm), or less than 0.001% (10ppm) by weight water, based on the total weight of the drug-loadedmicrobead composition.

Example 3

A drug-loaded microbead composition is prepared according to Example 1or Example 2. The drug-loaded microbead composition is then mixed withphysiologically buffered saline solution for a rehydration time of from5 minutes to 40 minutes to swell the microbeads and form an embolizationcomposition (A). The solution containing the swollen microbeads is theninjected into a patient.

To compare the in vitro doxorubicin release rates of the drug-loadedmicrobead composition prepared according to Example 1 or Example 2 tothose of other microbead compositions prepared from beads ofnon-degradable polymer materials, two comparative samples are preparedand injected into a patient. The first comparative sample (B) containsmicrobeads of biocompatible sulfonate-modified poly(vinyl alcohol)loaded with doxorubicin by adding the doxorubicin in an aqueous solutionto the unloaded microbeads and waiting about 24 hours before injection.The second comparative sample (C) contains microbeads of a sodiumacrylate alcohol copolymer formed by polymerization of vinyl acetate andmethyl acrylate and loaded with doxorubicin by adding the doxorubicin inan aqueous solution to the unloaded microbeads and waiting about 24hours before injection.

A comparison of the doxorubicin release rates of the three samples frominjection until 30 days after injection is provided in the FIGURE. Theembolization composition (A) prepared from the drug-loaded microbeadcomposition of Example 1 or Example 2 exhibits an initial burst ofdoxorubicin release over the first two days, during which approximately30% of the total doxorubicin content is released to the tissues of thepatient. The doxorubicin release continues through day 14, by which timeapproximately 70% of the total doxorubicin content of the microbeads hasbeen released, and further through day 30, by which time approximately90% to 100% of the total doxorubicin content of the microbeads has beenreleased.

In contrast, the first sample (B) releases approximately 20% of itsdoxorubicin immediately and by day 2 has released approximately 25% ofits doxorubicin content. After day 2, however, no further doxorubicin isreleased. Similarly, the second comparative sample (C) releasesapproximately 15% of its doxorubicin immediately and does not releaseany additional doxorubicin after the initial release.

Example 4

Liposomes including sorafenib incorporated within a lipid bilayer areprepared by subjecting a mixture of 37 mol. %poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), 12 mol. %dioleoyl phosphatidyl-serine (DOPS), 12 mol. % dioleoylphosphatidyl-ethanolamine (DOPE), 26 mol % cholesterol and 13 Mol %sorafenib in chloroform solvent to a centrifugation extrusion with a200-nm polycarbonate membrane. The size of the liposomes, as determinedby Dynamic Light Scattering (DLS), is 171.8±3.0 nm. Presence ofsorafenib in the final liposome preparation is verified from anabsorbance peak at about 270 nm, measured by UV-VIS spectroscopy.

Example 5

Liposomes including doxorubicin surrounded by a lipid bilayer areprepared by subjecting a mixture of PMPC, DOI'S, DOPE, and cholesterolin chloroform to a centrifugation extrusion with a 200-nm polycarbonatemembrane. The chloroform solvent is dried from the mixture. The lipidmixture is then rehydrated with an aqueous solution of HEPES, salt, anddoxorubicin, to form liposomes. Excess doxorubicin not incorporatedwithin the liposomes is removed by gel filtration.

Example 6

Ethosomes including sorafenib surrounded by a lipid bilayer are preparedby centrifugation extrusion with a 200-nm polycarbonate membrane of amixture of 42 mol. % PMPC, 14 mol. % DOPS, 14 mol. % DOPE, and 30 mol. %cholesterol in a 40% ethanol/water solution containing 2 mM sorafenib.Free sorafenib is separated from the ethosomes by size exclusionchromatography on Sephadex G-25. The sorafenib ethosomes have sizes234.4±4.5 nm determined by dynamic light scattering. Presence ofsorafenib in the final liposome preparation is verified from anabsorbance peak at about 270 nm, measured by UV-VIS spectroscopy.

Example 7

Liposomes including sorafenib within a lipid bilayer and doxorubicinsurrounded by the lipid are prepared by subjecting a mixture of 37 mol.% poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), 12 mol. %dioleoyl phosphatidyl-serine (DOPS), 12 mol. % dioleoylphosphatidyl-ethanolamine (DOPE), 26 mol. % cholesterol and 13 mol. %sorafenib in chloroform solvent to a centrifugation extrusion with a200-nm polycarbonate membrane. The solvent is dried from thelipid/sorafenib mixture. The lipid mixture is then rehydrated with anaqueous solution of HEPES, salt, and doxorubicin, to form liposomes.Excess doxorubicin not incorporated within the liposomes is removed bygel filtration.

Example 8

Vesicular agents such as the liposomes or ethosomes prepared accordingto any of Examples 4-7 are incorporated into microbeads of abiodegradable material by emulsion evaporation. An emulsion is preparedfrom an aqueous phase of deionized water and an emulsifier such aspolyvinyl alcohol and an organic phase containing the vesicular agentsand biodegradable polymer material.

An aqueous phase of 1% polyvinyl alcohol is prepared by stirring 8 gpolyvinyl alcohol (86-89% hydrolyzed) into 800 mL deionized water at 600rpm at 85° C. for 3-4 hours until the polyvinyl alcohol is fullydissolved.

An organic phase is prepared by adding to a 2-mL vial 150 mg celluloseacetate butyrate and 900 μL dichloromethane, sealing the vial, andshaking the vial until the cellulose acetate butyrate is dissolved. Tothe mixture, 100 μL of the vesicular agent in aqueous solution is added.

The organic phase and the aqueous phase are mixed by first adding to anappropriate vessel, such as a 250-mL flask, 100 mL of the aqueous phasethen adding the contents of the vial of organic phase into the flaskwhile stirring the aqueous phase in the flask at approximately 600 rpm.The flask is sealed with a lid, and stirring is continued for 30minutes. After the thirty minutes, the lid is removed and the contentsof the flask are stirred without the lid for an additional 2 hours. Thestirring is then stopped, and the microbeads that have formed in thesolution are allowed to settle.

The microbeads are then transferred by pipette to centrifuge tubes, andthe tubes are centrifuged at 12000 rpm or greater for at least 2.5minutes. The supernatant is then removed by transfer pipette, and themicrobeads are washed by refilling the tubes a with deionized orfiltered water and centrifuging again at 12000 rpm or greater for atleast 2.5 minutes. The washing process is repeated from one to fivetimes. The resulting microbeads have sizes from about 20 μm to about 350μm.

The centrifuged microbeads are vacuum filtered through 200-micron meshfilters into a vacuum flask. The filtered microbeads in the flask havesizes from about 20 μm to about 200 The filtered microbeads in the flaskare then further vacuum filtered through a 100-micron mesh filter. Themicrobeads remaining on the filter are retained and have sizes fromabout 100 μm to about 200 These microbeads are rinsed with filteredwater then are allowed to dry in air for 24 hours. The air-driedmicrobeads then may be transferred to a suitable container or vial.

Items Listing

Embodiments of the present disclosure include at least following Items,which are not intended to limit the scope of the disclosure as a wholeor the appended claims.

Item A1: A drug-loaded microbead composition comprising: microbeadscomprising a water-swellable polymer material; and a complex comprisinga carrier and a therapeutic agent chemically bonded to the carrier,wherein: the complex is embedded within the polymer material; and thedrug-loaded microbead composition has a water content of less than 1% byweight, based on the total weight of the drug-loaded microbeadcomposition.

Item A2: The drug-loaded microbead composition of Item A1, wherein thewater-swellable polymer material is a biodegradable polymer materialchosen from poly(-hydroxybutyrate), methacrylated hyaluronic acids,chitosan-alginate, poly(N-isopropylacrylamide) copolymers,poly(N-isopropylacrylamide)-alginates,poly(N-isopropylacrylamide)-peptides,poly(N-isopropylacrylamide)-α-acryloyloxy-β,β-dimethyl-γ-butyrolactone-hydrophilicJeffamine, poly(N-isopropylacrylamide)-poly(ethylene glycol)diacrylate-pentaerythritol tetrakis(3-mercaptopropionate), derivativesof any of the foregoing, or combinations of any of the foregoing.

Item A3: The drug-loaded microbead composition of Item A1 or A2, whereinthe therapeutic agent is chosen from doxorubicin, sorafenib, vandetanib,nivolumab, ipilimumab, regorafenib, irinotecan, epirubicin, pirarubicin,5-fluorouracil, cisplatin, floxuridine, mitomycin C, derivatives of anyof the foregoing, prodrugs of any of the foregoing, therapeuticallyacceptable salts of any of the foregoing, or combinations of any of theforegoing.

Item A4: The drug-loaded microbead composition of any one of thepreceding Items, wherein the therapeutic agent is not chemically bondedto the water-swellable polymer material.

Item A5: The drug-loaded microbead composition of any one of thepreceding Items, wherein: the water-swellable polymer material comprisesa polymer matrix shell encapsulating a particle core; and the complex isdisposed within the particle core.

Item A6: The drug-loaded microbead composition of any one of thepreceding Items, wherein the carrier is embedded within aninterpenetrating network of the water-swellable polymer material of thedrug-loaded microbead.

Item A7: The drug-loaded microbead composition of any one of thepreceding Items, wherein the carrier is chosen from liposomes, polymericmicelles, Pluronics, polycaprolactone-b-methoxy-PEG, poly(asparticacid)-b-PEG, poly(benzyl-L-glutamate)-b-PEG,poly(D,L-lactide)-b-methoxy-PEG, poly(β-benzyl-L-aspartate)-b-PEG), orsodium dextran sulfate.

Item A8: The drug-loaded microbead composition of Item A6, wherein thecarrier comprises a sodium dextran sulfate.

Item A9: The drug-loaded microbead composition of Item A8, wherein thesodium dextran sulfate has a molecular weight of from about 40 kDa toabout 500 kDa.

Item A10: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises liposomes.

Item A11: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises polymeric micelles.

Item A12: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises Pluronics.

Item A13: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises polycaprolactone-b-methoxy-PEG.

Item A14: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises poly(aspartic acid)-b-PEG.

Item A15: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises poly(benzyl-L-glutamate)-b-PEG.

Item A16: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises poly(D,L-lactide)-b-methoxy-PEG.

Item A17: The drug-loaded microbead composition of Item A7, wherein thecarrier comprises poly(β-benzyl-L-aspartate)-b-PEG).

Item A18: The drug-loaded microbead composition of any one of thepreceding Items, wherein the drug-loaded microbeads are substantiallyspherical.

Item A19: The drug-loaded microbead composition of any one of thepreceding Items, wherein the drug-loaded microbeads have diameters fromabout 40 μm to about 800 μm.

Item A20: The drug-loaded microbead composition of any one of thepreceding Items, wherein individual microbeads of the drug loadedmicrobead composition comprise, based on the total weight of theindividual microbeads: from about 30% by weight to about 70% by weightwater-swellable polymer material; from about 1% by weight to about 25%by weight therapeutic agent; from about 1% by weight to about 40% byweight carrier; and less than 1% by weight water.

Item A21: A method for preparing a drug-loaded microbead composition,the method comprising: dissolving a carrier and a therapeutic agent inan aqueous solvent to form an initial mixture, the initial mixturecomprising a complex of the carrier and the therapeutic agent; combiningthe initial mixture with a first solvent composition in which thetherapeutic agent is less soluble than in the aqueous solvent to form asecond mixture; stirring the second mixture rapidly to form particles ofthe complex in the second mixture; combining the particles of thecomplex with a hydrogel of a water-swellable polymer material to form ahydrogel mixture; combining the hydrogel mixture with a second solventcomposition to form a synthesis mixture; stirring the synthesis mixturerapidly to form drug-loaded microbeads in the synthesis mixture;recovering the drug-loaded microbeads from the synthesis mixture; andremoving water from the drug-loaded microbeads to form a drug-loadedmicrobead composition having a water content of less than 1% by weight,based on the total weight of the drug-loaded microbeads in thedrug-loaded microbead composition.

Item A22: The method of Item A21, further comprising filtering thesecond mixture to recover the particles of the complex before combiningthe particles of the complex with the hydrogel solution.

Item A23: The method of Item A21 or A22, wherein the removing watercomprises lyophilizing the drug-loaded microbeads.

Item A24: The method of Item A21 or A22, wherein: the removing watercomprises lyophilizing the drug-loaded microbeads; and drying thelyophilized drug-loaded microbeads by air, temperature variation,vacuum, or any combination thereof.

Item A25: The method of any one of Items A21-A24, wherein: thedrug-loaded microbeads have an average synthesized volume when recoveredfrom the synthesis mixture and an average final volume after theremoving of water; and the average final volume is from about 10% toabout 75% of the average synthesized volume.

Item A26: A method for preparing a drug-loaded microbead composition,the method comprising: dissolving a carrier and a therapeutic agent inan aqueous solvent to form an initial mixture, the initial mixturecomprising a complex of the carrier and the therapeutic agent; combiningthe initial mixture with a hydrogel solution of a water-swellablepolymer material to form a hydrogel mixture; combining the hydrogelmixture with a solvent composition to form a synthesis mixture; stirringthe synthesis mixture rapidly to form drug-loaded microbeads in thesynthesis mixture; recovering the drug-loaded microbeads from thesynthesis mixture; and drying the drug-loaded microbeads to form adrug-loaded microbead composition having a water content of less than 1%by weight, based on the total weight of the drug-loaded microbeads inthe drug-loaded microbead composition.

Item A27: The method of Item A26, wherein the removing water compriseslyophilizing the drug-loaded microbeads.

Item A28: The method of Item A26, wherein: the removing water compriseslyophilizing the drug-loaded microbeads; and drying the lyophilizeddrug-loaded microbeads in air, heat, vacuum, or any combination thereof.

Item A29: The method of any one of Items A26-A28, wherein thedrug-loaded microbeads have an average synthesized volume when recoveredfrom the synthesis mixture and an average final volume after theremoving of water; and the average final volume is from about 10% toabout 75% of the average synthesized volume.

Item A30: An embolization composition comprising: a drug-loadedmicrobead composition according to any of Items A1-A20, or a drug-loadedmicrobead composition made by the method according to any of ItemsA21-A25, or a drug-loaded microbead composition made by the methodaccording to any of Items A26-A29; and an amount of an aqueous solutionsufficient to cause the drug-loaded microbeads of the drug-loadedmicrobead composition to swell, whereby the drug-loaded microbeads afterswelling comprise a water content of from 50% by weight to 95% byweight, based on the total weight of the drug-loaded microbeads.

Item A31: The embolization composition of Item A30, wherein the aqueoussolution is a physiologically buffered saline solution.

Item A32: The embolization composition of Item A30 or A31, whereinswollen individual drug-loaded microbeads of the embolizationcomposition comprise, based on the total weight of the swollenindividual drug-loaded microbeads: from about 3% by weight to about 10%by weight water-swellable polymer material; from about 0.1% by weight toabout 2.5% by weight therapeutic agent; from about 0.1% by weight toabout 4.0% by weight carrier; and from about 50% by weight to about 96%by weight water.

Item A33: A method for preparing an embolization composition, the methodcomprising: adding a drug-loaded microbead composition according to anyof Items A1-A20, or a drug-loaded microbead composition made accordingto the method of any one of Items A21-A25, or a drug-loaded microbeadcomposition made according to the method of any one of Items A26-A29, toan amount of an aqueous solution sufficient to cause the drug-loadedmicrobeads of the drug-loaded microbead composition to swell, wherebythe drug-loaded microbeads after swelling comprise a water content offrom 50% by weight to 95% by weight, based on the total weight of thedrug-loaded microbeads, to form an injection-ready solution; and loadingthe injection-ready solution into an injection device.

Item A34: The method of Item A33, further comprising allowing thedrug-loaded microbeads to swell for a rehydration time of from about 5minutes to about 60 minutes before loading the injection-ready solutioninto the injection device.

Item A35: The method of Item A33 or A34, wherein the aqueous solution isa physiologically buffered saline solution.

Item A36: A method for treating a disease, the method comprising:providing intravenously, to a subject in need of embolization therapy,an embolization composition according to any one of Items A30-A32.

Item A37: The method of Item A36, wherein: after the embolizationcomposition is provided intravenously, at least a portion of thedrug-loaded microbeads flow through vasculature of the subject to anembolization site and restrict blood flow through the embolization site;and the drug-loaded microbeads release to tissue at the embolizationsite over the course of a release period at least 90% by weight of thetherapeutic agent, based on an initial amount of therapeutic agentembedded within the microbeads before the embolization composition isprovided.

Item A38: The method of Item A37, wherein the release to the tissue atthe embolization site includes an initial burst of release, during whichof at least 10% by weight of the therapeutic agent, based on the initialamount.

Item A39: The method of Item A37 or A38, wherein the drug-loadedmicrobeads provide a sustained release of the therapeutic agent to thetissue over the release period.

Item A40: The method of any one of Items A37-A39, wherein the releaseperiod is at least 14 days.

Item A41: The method of any one of Items A37-A38, wherein the releaseperiod is at least 28 days.

Item A42: The method of any one of Items A36-A41, wherein the disease isa cancer.

Item A43: The method of Item A42, wherein the cancer is hepatocellularcarcinoma, liver cancer, prostate cancer, or breast cancer.

Item B1: A drug-loaded microbead composition comprising: a plurality ofindividual microbeads of a biodegradable material; a plurality ofindividual vesicular agents contained within the individual microbeadsor associated with the biodegradable material of the individualmicrobeads through ionic or non-covalent interaction, the vesicularagents comprising at least one lipid bilayer surrounding a vesicularcore; and a first therapeutic agent contained within the individualvesicular agents or associated with the individual vesicular agents byionic or non-covalent interaction; and a second therapeutic agentdifferent from the first therapeutic agent and contained within theindividual microbeads or associated with the individual microbeadsthrough ionic or non-covalent interaction.

Item B2: The drug-loaded microbead composition of Item B1, wherein thevesicular agents comprise liposomes or ethosomes.

Item B3: The drug-loaded microbead composition of Item B1 or B2, whereinthe second therapeutic agent is contained within the individualvesicular agents or associated with individual vesicular agents throughionic or non-covalent interaction.

Item B4: The drug-loaded microbead composition of any of the precedingItems B1 to B3, wherein the first therapeutic agent is hydrophilic andthe second therapeutic agent is hydrophobic.

Item B5: The drug-loaded microbead composition of Item B4, wherein thefirst therapeutic agent is encapsulated within the vesicular core of theindividual vesicular agents and the second therapeutic agent iscontained within the lipid bilayer of the individual vesicular agents.

Item B6: The drug-loaded microbead composition of Item B4, wherein thesecond therapeutic agent is not contained within the individualvesicular agents and is associated by ionic or non-covalent interactionwith the biodegradable material of the individual microbeads.

Item B7: The drug-loaded microbead composition of any of the precedingItems B1 to B6, wherein the first therapeutic agent and the secondtherapeutic agent are independently chosen from doxorubicin,bevacizumab, bortezomib, imatinib, seliciclib, ceritinib, everolimus,paclitaxel, sorafenib, irinotecan, idarubicin, cisplatin, andcombinations thereof.

Item B8: The drug-loaded microbead composition of any of the precedingItems B1 to B7, wherein the biodegradable material is a biodegradablepolymer chosen from poly(lactide-co-glycolide) (PLGA), polylactide(PLA), polyglycolide (PGA), polycaprolactone (PCL),polyhydroxyalkanoates, poly-R-3-hydroxybutyrate (poly3HB),poly-R-3-hydroxybutyrate-co-R-3-hydroxyvalerate (poly(3HB-co-3HV)),poly-R-3-hydroxybutyrate-co-4-hydroxybutyrate (poly(3HB-co-4HB)),poly-R-3 hydroxyoctanoate-co-R-3-hydroxyhexanoate (poly(3HO-co-3HH)),poly-3-hydroxypropionate (poly(3HP)), poly-4-hydroxybutyrate(poly(4HB)), poly-5-hydroxybutyrate (poly(5HB)), poly-6-hydroxybutyrate(poly(6HB)), poly(propylene fumarate), poly(butylene succinate),poly(p-dioxanone, polyacetals, poly(ortho esters), polycarbonates,chitosan, hydroxybutyric acids, polyanhydrides and polyesters,polyphosphazenes, polyphosphoesters, lipodisq, celluloses, modifiedcelluloses, proteins and poly(amino acids), polyethers, and co-polymersof the foregoing.

Item B9: The drug-loaded microbead composition of any of the precedingItems B1 to B8, wherein the biodegradable material is a cellulose.

Item B10: The drug-loaded microbead composition of any of the precedingItems B1 to B9, wherein the biodegradable material is a modifiedcellulose comprising cellulose acetate butyrate.

Item B11: The drug-loaded microbead composition of any of Items B1 toB7, wherein the biodegradable material is a lipid chosen from tricaprin,trilaurin, trimyristin, tripalmitin, tristearin, hydrogenatedcoco-glycerides, glyceryl monostearate, glyceryl behenate, glycerylpalmitostearate, glyceryl caprate, cetyl palmitate, stearic acid,palmitic acid, decanoic acid, behenic acid, beeswax, carnauba wax, cacaobutter, and combinations thereof.

Item B12: The drug-loaded microbead composition of Item B1, wherein: theindividual vesicular agents comprise liposomes; the first therapeuticagent is contained within the lipid bilayer of the individual vesicularagents; the second therapeutic agent is not contained within theindividual vesicular agents and is associated by ionic or non-covalentinteraction with the biodegradable material of the individualmicrobeads; and the biodegradable material is a cellulose.

Item B13: The drug-loaded microbead composition of Item B1, wherein: theindividual vesicular agents comprise liposomes; the first therapeuticagent is contained within the lipid bilayer of the individual vesicularagents; the second therapeutic agent is encapsulated within thevesicular core of the individual vesicular agents; the first therapeuticagent is sorafenib; the second therapeutic agent is doxorubicin; and thebiodegradable material is a cellulose.

Item B14: The drug-loaded microbead composition of Item B1, wherein: theindividual vesicular agents comprise ethosomes; the first therapeuticagent is encapsulated within the vesicular core of the individualvesicular agents; the second therapeutic agent is not contained withinthe individual vesicular agents and is associated by ionic ornon-covalent interaction with the biodegradable material; the firsttherapeutic agent is sorafenib; the second therapeutic agent isdoxorubicin; and the biodegradable material is a cellulose.

Item B15: The drug-loaded microbead composition of any of the precedingItems B1 to B14, wherein the at least one lipid bilayer comprisespoly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), dioleoylphosphatidylserine (DOPS), dioleoyl phosphatidylethanolamine (DOPE), andcholesterol.

Item C1: A drug-loaded microbead composition according to any of ItemsB1 to B15, comprising as a further attribute any feature of anydrug-loaded microbead composition according to any of Items A1 to A20.

Item C2: A drug-loaded microbead composition according to any of ItemsA1 to A20, comprising as a further attribute any feature of anydrug-loaded microbead composition according to any of Items B1 to B15.

Item C3: A drug-loaded microbead composition comprising a combination ormixture of microbeads from a drug-loaded microbead composition accordingto any of Items A1 to A20 and microbeads from a drug-loaded microbeadcomposition according to any of Items B1 to B15.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which the invention belongs. The terminology used in thedescription herein is for describing particular embodiments only and isnot intended to be limiting. As used in the specification and appendedclaims, the singular forms “a,” “an,” and “the” are intended to includethe plural forms as well, unless the context clearly indicatesotherwise.

Unless otherwise indicated, all numbers expressing quantities ofingredients, properties such as molecular weight, reaction conditions,and so forth as used in the specification and claims are to beunderstood as being modified in all instances by the term “about.”Accordingly, unless otherwise indicated, the numerical properties setforth in the specification and claims are approximations that may varydepending on the desired properties sought to be obtained in embodimentsof the present invention. Notwithstanding that the numerical ranges andparameters setting forth the broad scope of the invention areapproximations, the numerical values set forth in the specific examplesare reported as precisely as possible. One of ordinary skill in the artwill understand that any numerical values inherently contain certainerrors attributable to the measurement techniques used to ascertain thevalues.

It is noted that terms like “preferably,” “commonly,” and “typically”are not utilized herein to limit the scope of the claims or to implythat certain features are critical, essential, or even important to thestructure or function of the claimed invention. Rather, these terms aremerely intended to highlight alternative or additional features that mayor may not be utilized in a particular embodiment of the presentinvention.

For the purposes of describing and defining the present invention it isnoted that the term “substantially” is used herein to represent theinherent degree of uncertainty that may be attributed to anyquantitative comparison, value, measurement, or other representation.The term “substantially” is used herein also to represent the degree bywhich a quantitative representation may vary from a stated referencewithout resulting in a change in the basic function of the subjectmatter at issue. As such, it is used to represent the inherent degree ofuncertainty that may be attributed to any quantitative comparison,value, measurement, or other representation, referring to an arrangementof elements or features that, while in theory would be expected toexhibit exact correspondence or behavior, may in practice embodysomething less than exact.

It will be apparent to those skilled in the art that variousmodifications and variations can be made to the embodiments describedherein without departing from the spirit and scope of the claimedsubject matter. Thus it is intended that the specification cover themodifications and variations of the various embodiments described hereinprovided such modification and variations come within the scope of theappended claims and their equivalents.

The invention claimed is:
 1. A drug-loaded microbead compositioncomprising: a plurality of individual microbeads of a biodegradablematerial; a plurality of individual vesicular agents contained withinthe individual microbeads or associated with the biodegradable materialof the individual microbeads through ionic or non-covalent interaction,the vesicular agents comprising at least one lipid bilayer comprising aphospholipid and surrounding a vesicular core, wherein the vesicularagents comprise liposomes or ethosomes; and a first therapeutic agentcontained within the individual vesicular agents or associated with theindividual vesicular agents by ionic or non-covalent interaction; and asecond therapeutic agent different from the first therapeutic agent andcontained within the individual microbeads or associated with theindividual microbeads through ionic or non-covalent interaction.
 2. Thedrug-loaded microbead composition of claim 1, wherein the secondtherapeutic agent is contained within the individual vesicular agents orassociated with individual vesicular agents through ionic ornon-covalent interaction.
 3. The drug-loaded microbead composition ofclaim 1, wherein the first therapeutic agent is hydrophilic and thesecond therapeutic agent is hydrophobic.
 4. The drug-loaded microbeadcomposition of claim 3, wherein the first therapeutic agent isencapsulated within the vesicular core of the individual vesicularagents and the second therapeutic agent is contained within the lipidbilayer of the individual vesicular agents.
 5. The drug-loaded microbeadcomposition of claim 3, wherein the second therapeutic agent is notcontained within the individual vesicular agents and is associated byionic or non-covalent interaction with the biodegradable material of theindividual microbeads.
 6. The drug-loaded microbead composition of claim1, wherein the first therapeutic agent and the second therapeutic agentare independently chosen from doxorubicin, bevacizumab, bortezomib,imatinib, seliciclib, ceritinib, everolimus, paclitaxel, sorafenib,irinotecan, idarubicin, cisplatin, and combinations thereof.
 7. Thedrug-loaded microbead composition of claim 1, wherein the biodegradablematerial is selected from the group consisting of biodegradable polymersand lipids, wherein: the biodegradable polymers are chosen frompoly(lactide-co-glycolide) (PLGA), polylactide(PLA), polyglycolide(PGA), polycaprolactone (PCL), polyhydroxyalkanoates,poly-R-3-hydroxybutyrate (poly3HB),poly-R-3-hydroxybutyrate-co-R-3-hydroxyvalerate (poly(3RB-co-3HV)),poly-R-3-hydroxybutyrate-co-4-hydroxybutyrate (poly(3HB-co-4HB)),poly-R-3-hydroxyoctanoate-co-R-3-hydroxyhexanoate (poly(3HO-co-3HH)),poly-3-hydroxypropionate (poly(3HP)), poly-4-hydroxybutyrate(poly(4HB)), poly-5-hydroxybutyrate (poly(5H-1B)),poly-6-hydroxybutyrate (poly(6HB)), polypropylene fumarate),poly(butylene succinate), poly(p-dioxanone, polyacetals, poly(orthoesters), polycarbonates, chitosan, hydroxybutyric acids, polyanhydrides,polyphosphazenes, polyphosphoesters, celluloses or modified celluloses,proteins, poly(amino acids), polyethers, and co-polymers of theforegoing; and the lipids are chosen from tricaprin, trilaurin,trimyristin, tripalmitin, tristearin, hydrogenated coco-glycerides,glyceryl monostearate, glyceryl behenate, glyceryl palmitostearate,glyceryl caprate, cetyl palmitate, stearic acid, palmitic acid, decanoicacid, behenic acid, beeswax, carnauba wax, cacao butter, andcombinations thereof.
 8. The drug-loaded microbead composition of claim1, wherein: the individual vesicular agents comprise liposomes; thefirst therapeutic agent is contained within the lipid bilayer of theindividual vesicular agents; the second therapeutic agent is notcontained within the individual vesicular agents and is associated byionic or non-covalent interaction with the biodegradable material of theindividual microbeads; and the biodegradable material is a cellulose. 9.The drug-loaded microbead composition of claim 1, wherein: theindividual vesicular agents comprise liposomes; the first therapeuticagent is contained within the lipid bilayer of the individual vesicularagents; the second therapeutic agent is encapsulated within thevesicular core of the individual vesicular agents; the first therapeuticagent is sorafenib; the second therapeutic agent is doxorubicin; and thebiodegradable material is a cellulose.
 10. The drug-loaded microbeadcomposition of claim 1, wherein: the individual vesicular agentscomprise ethosomes; the first therapeutic agent is encapsulated withinthe vesicular core of the individual vesicular agents; the secondtherapeutic agent is not contained within the individual vesicularagents and is associated by ionic or non-covalent interaction with thebiodegradable material; the first therapeutic agent is sorafenib; thesecond therapeutic agent is doxorubicin; and the biodegradable materialis a cellulose.
 11. The drug-loaded microbead composition of claim 1,wherein the at least one lipid bilayer comprises a combination ofpoly(2-methacryloyloxyethyl phosphorylcholine) (PMPC), dioleoylphosphatidylserine (DOPS), dioleoyl phosphatidylethanolamine (DOPE), andcholesterol.